GcatWiki - User contributions [en]

Bio-Math Connections January - May 2010

2010-01-29T18:29:18Z

Cdavis21:

Write in which project you will present, your name, and which campus you represent. For example:

* MOWestern/Davidson 2009 project:: Malcolm Campbell:: Davidson
* Cambridge 2009 project:: Michael Rydberg, Nitya Rao, Erin Feeney:: Davidson
* U.C. Berkeley 2009 project:: Anvi Raina, Steph Meador, Linda Kleist:: Davidson
* S.J.T.U. Shanghai 2009 project:: Yihharn Hwang, Stephen Streb, Shashank Suresh:: Davidson
* Stanford 2009 project:: Kris Hendershot, Garrett Smith, Tom Shuman:: Davidson
* NCTU Formosa/WetLab 2009 project:: Clif Davis:: Missouri Western

IGEM 2009 Project

2009-07-02T12:53:58Z

Cdavis21:

Biology Based:
#[[Which reporters are we going to use?]]
#[[What naming system are we going to use for the suppressor tRNAs?]]
#[[How do you build the tRNA construct?]]
#[[How are we going to build the 5-mer BioBricks?]]

[http://gcat.davidson.edu/GcatWiki/images/2/2c/Stop_Codons_in_LCs.doc Using Stop Codons to Truncate Translation] 
[http://gcat.davidson.edu/GcatWiki/images/8/81/D5merSequences.doc Davidson ATG+5mer BioBrick Sequences] 
[http://gcat.davidson.edu/GcatWiki/images/4/47/FML_CODING_SEQUENCE_2.doc Frameshift Mutation Leader PCR primers] 
[http://www.staff.uni-bayreuth.de/~btc914/search/index.html Align E. coli tRNA sequences] 
[http://media.pearsoncmg.com/bc/bc_campbell_genomics_2/medialib/jmol/trna/index.html Compare 2 tRNA structures] 

'''Team Progress Table'''

{| border="1" cellpadding="2"
!width="120"|Student Name
!width="30"|5mer Codon
!width="40"|Anticodon Sequence
!width="40"| [http://gcat.davidson.edu/GcatWiki/index.php/What_naming_system_are_we_going_to_use_for_the_suppressor_tRNAs%3F Codon Short Name]
!width="150"|tRNA Status
!width="150"|5mer+XFP Status
!width="150"|5mer+Drug resistance Status
|-
| Olivia Ho-Shing || CCAUC || GUGAUCCAA-9 || Pro4 || 2 clones being resequenced || 2 5mer+RFPs sequencing || 5 5mer+Tets sequencing
|-
| Olivia Ho-Shing || CCAUC || UUUGAUGGAG-10 || Pro5 || 1 clone being resequenced || 2 5mer+RFPs sequencing || 5 5mer+Tets sequencing
|-
| Romina Clemente|| CUAGU || UUACUAGAC-9 || Leu4|| '''1 clone 100% match''', putting pBAD upstream || 9 5mer+RFPs sequencing || 5 5mer+Tets sequencing
|-
| Shamita Punjabi|| CCACU || CUAGUGGAC-9 || Pro3|| 2 clones being resequenced || 6 5mer+RFPs sequencing || 5 5mer+Tets sequencing
|-
| Leland Taylor || CCCUC || CUGAGGGUC-9 || Pro6 || '''1 clone 100% match''', putting pBAD upstream || 7 5mer+RFPs sequencing || 7 5mer+Tets sequencing
|-
| Alyndria Thompson|| CGGUC || UUGACCGAC-9 || Arg2 || 5 clones being sequenced || 4 5mer+RFPs sequencing || 6 5mer+Tets sequencing
|-
| Clif Davis|| 5mer codon || Anticodon Sequence || Leu3|| tRNA Status || 5mer+RFP Status || 2 5mer+CAT sequenced-99% Match with a Conservative Mutation
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
|}

Math Based: (Davidson) We have posted a Mathematica notebook on the Wiggio. Basically we are at a point where we can choose the number of variables, SAT number(2-SAT, 3-SAT, etc), and the number of clauses to use at a time and produce a table of inputs as the rows and clauses as the columns, with a 1 if the input satisfies the clause and a 0 if the input fails to satisfy the clause. In addition, we can produce a "Super Table," which lists all the SAT problems given the number of clauses in conjunctive normal form and displays how many of the clauses in a given problem each input satisfies. Feel free to view the posted Mathematica notebook; there are some instructions within the notebook itself. When you open it, there will be a window which asks about dynamic content; be sure to click "Enable Dynamic."

IGEM 2009 Project

2009-06-29T19:58:14Z

Cdavis21:

Biology Based:
#[[Which reporters are we going to use?]]
#[[What naming system are we going to use for the suppressor tRNAs?]]
#[[How do you build the tRNA construct?]]
#[[How are we going to build the 5-mer BioBricks?]]

[http://gcat.davidson.edu/GcatWiki/images/2/2c/Stop_Codons_in_LCs.doc Using Stop Codons to Truncate Translation] 
[http://gcat.davidson.edu/GcatWiki/images/8/81/D5merSequences.doc Davidson ATG+5mer BioBrick Sequences] 
[http://gcat.davidson.edu/GcatWiki/images/4/47/FML_CODING_SEQUENCE_2.doc Frameshift Mutation Leader PCR primers] 
[http://www.staff.uni-bayreuth.de/~btc914/search/index.html Align E. coli tRNA sequences] 
[http://media.pearsoncmg.com/bc/bc_campbell_genomics_2/medialib/jmol/trna/index.html Compare 2 tRNA structures] 

'''Team Progress Table'''

{| border="1" cellpadding="2"
!width="120"|Student Name
!width="30"|5mer Codon
!width="40"|Anticodon Sequence
!width="40"| [http://gcat.davidson.edu/GcatWiki/index.php/What_naming_system_are_we_going_to_use_for_the_suppressor_tRNAs%3F Codon Short Name]
!width="150"|tRNA Status
!width="150"|5mer+XFP Status
!width="150"|5mer+Drug resistance Status
|-
| Olivia Ho-Shing || CCAUC || GUGAUCCAA-9 || Pro4 || 2 clones being resequenced || 2 5mer+RFPs sequencing || 5 5mer+Tets sequencing
|-
| Olivia Ho-Shing || CCAUC || UUUGAUGGAG-10 || Pro5 || 1 clone being resequenced || 2 5mer+RFPs sequencing || 5 5mer+Tets sequencing
|-
| Romina Clemente|| CUAGU || UUACUAGAC-9 || Leu4|| '''1 clone 100% match''', putting pBAD upstream || 9 5mer+RFPs sequencing || 5 5mer+Tets sequencing
|-
| Shamita Punjabi|| CCACU || CUAGUGGAC-9 || Pro3|| 2 clones being re-sequenced || 6 5mer+RFPs sequencing || 5 5mer+Tets sequencing
|-
| Leland Taylor || CCCUC || CUGAGGGUC-9 || Pro6 || '''1 clone 100% match''', putting pBAD upstream || 7 5mer+RFPs sequencing || 7 5mer+Tets sequencing
|-
| Alyndria Thompson|| CGGUC || UUGACCGAC-9 || Arg2 || 5 clones being sequenced || 4 5mer+RFPs sequencing || 6 5mer+Tets sequencing
|-
| Clif Davis|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 2 5mer+CAT sequencing
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
|}

Math Based: (Davidson) We have posted a Mathematica notebook on the Wiggio. Basically we are at a point where we can choose the number of variables, SAT number(2-SAT, 3-SAT, etc), and the number of clauses to use at a time and produce a table of inputs as the rows and clauses as the columns, with a 1 if the input satisfies the clause and a 0 if the input fails to satisfy the clause. In addition, we can produce a "Super Table," which lists all the SAT problems given the number of clauses in conjunctive normal form and displays how many of the clauses in a given problem each input satisfies. Feel free to view the posted Mathematica notebook; there are some instructions within the notebook itself. When you open it, there will be a window which asks about dynamic content; be sure to click "Enable Dynamic."

IGEM 2009 Project

2009-06-29T19:56:20Z

Cdavis21:

Biology Based:
#[[Which reporters are we going to use?]]
#[[What naming system are we going to use for the suppressor tRNAs?]]
#[[How do you build the tRNA construct?]]
#[[How are we going to build the 5-mer BioBricks?]]

[http://gcat.davidson.edu/GcatWiki/images/2/2c/Stop_Codons_in_LCs.doc Using Stop Codons to Truncate Translation] 
[http://gcat.davidson.edu/GcatWiki/images/8/81/D5merSequences.doc Davidson ATG+5mer BioBrick Sequences] 
[http://gcat.davidson.edu/GcatWiki/images/4/47/FML_CODING_SEQUENCE_2.doc Frameshift Mutation Leader PCR primers] 
[http://www.staff.uni-bayreuth.de/~btc914/search/index.html Align E. coli tRNA sequences] 
[http://media.pearsoncmg.com/bc/bc_campbell_genomics_2/medialib/jmol/trna/index.html Compare 2 tRNA structures] 

'''Team Progress Table'''

{| border="1" cellpadding="2"
!width="120"|Student Name
!width="30"|5mer Codon
!width="40"|Anticodon Sequence
!width="40"| [http://gcat.davidson.edu/GcatWiki/index.php/What_naming_system_are_we_going_to_use_for_the_suppressor_tRNAs%3F Codon Short Name]
!width="150"|tRNA Status
!width="150"|5mer+XFP Status
!width="150"|5mer+Drug resistance Status
|-
| Olivia Ho-Shing || CCAUC || GUGAUCCAA-9 || Pro4 || 2 clones being resequenced || 2 5mer+RFPs sequencing || 5 5mer+Tets sequencing
|-
| Olivia Ho-Shing || CCAUC || UUUGAUGGAG-10 || Pro5 || 1 clone being resequenced || 2 5mer+RFPs sequencing || 5 5mer+Tets sequencing
|-
| Romina Clemente|| CUAGU || UUACUAGAC-9 || Leu4|| '''1 clone 100% match''', putting pBAD upstream || 9 5mer+RFPs sequencing || 5 5mer+Tets sequencing
|-
| Shamita Punjabi|| CCACU || CUAGUGGAC-9 || Pro3|| 2 clones being re-sequenced || 6 5mer+RFPs sequencing || 5 5mer+Tets sequencing
|-
| Leland Taylor || CCCUC || CUGAGGGUC-9 || Pro6 || '''1 clone 100% match''', putting pBAD upstream || 7 5mer+RFPs sequencing || 7 5mer+Tets sequencing
|-
| Alyndria Thompson|| CGGUC || UUGACCGAC-9 || Arg2 || 5 clones being sequenced || 4 5mer+RFPs sequencing || 6 5mer+Tets sequencing
|-
| Clif Davis|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 2 5mer+CAT Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
| Student Name|| 5mer codon || Anticodon Sequence || Codon Short Name|| tRNA Status || 5mer+RFP Status || 5mer+Tet Resistance Status
|-
|}

Math Based: (Davidson) We have posted a Mathematica notebook on the Wiggio. Basically we are at a point where we can choose the number of variables, SAT number(2-SAT, 3-SAT, etc), and the number of clauses to use at a time and produce a table of inputs as the rows and clauses as the columns, with a 1 if the input satisfies the clause and a 0 if the input fails to satisfy the clause. In addition, we can produce a "Super Table," which lists all the SAT problems given the number of clauses in conjunctive normal form and displays how many of the clauses in a given problem each input satisfies. Feel free to view the posted Mathematica notebook; there are some instructions within the notebook itself. When you open it, there will be a window which asks about dynamic content; be sure to click "Enable Dynamic."

Can we solve a 3-SAT problem with supressor logic?

2009-05-27T14:55:17Z

Cdavis21: /* Things we need to learn about */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif) I don't know how to post the powerpoint on here.
[http://gcat.davidson.edu/GcatWiki/images/d/d1/3SAT_ppt-1-.ppt Bryce and Clif's Version]

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

[http://gcat.davidson.edu/GcatWiki/images/e/e0/PNAS-2006-Rodriguez-8650-5.pdf Rodriguez, Lester, Dougherty]

[http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6T36-4HM7N2R-G&_user=2665120&_rdoc=1&_fmt=&_orig=search&_sort=d&view=c&_acct=C000058476&_version=1&_urlVersion=0&_userid=2665120&md5=d2dda7b99095c256fd459f2f6c82b493 3 Suppressor tRNAs in a single protein, in vitro]

Incorporating unnatural amino acids using frameshift suppression. They used three in one eukaryotic cell.
<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-27T14:53:50Z

Cdavis21: /* Things we need to learn about */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif) I don't know how to post the powerpoint on here.
[http://gcat.davidson.edu/GcatWiki/images/d/d1/3SAT_ppt-1-.ppt Bryce and Clif's Version]

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

[http://gcat.davidson.edu/GcatWiki/images/e/e0/PNAS-2006-Rodriguez-8650-5.pdf Rodriguez, Lester, Dougherty]

[http://www.sciencedirect.com/science?_ob=ArticleURL&_udi=B6T36-4HM7N2R-G&_user=2665120&_rdoc=1&_fmt=&_orig=search&_sort=d&view=c&_acct=C000058476&_version=1&_urlVersion=0&_userid=2665120&md5=d2dda7b99095c256fd459f2f6c82b493]

Incorporating unnatural amino acids using frameshift suppression. They used three in one eukaryotic cell.
<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T18:19:06Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif) I don't know how to post the powerpoint on here.
[http://gcat.davidson.edu/GcatWiki/index.php/Image:3SAT_ppt-1-.ppt Bryce and Clif's Version]

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T18:16:55Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif) I don't know how to post the powerpoint on here.
[http://gcat.davidson.edu/GcatWiki/index.php/Image:3SAT_ppt-1-.ppt]

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T18:13:11Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif) I don't know how to post the powerpoint on here.
[http://gcat.davidson.edu/GcatWiki/images/6/6f/3SAT_ppt-1-.ppt]

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T18:07:51Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif) I don't know how to post the powerpoint on here.

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T18:07:39Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif)I don't know how to post the powerpoint on here.

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T18:07:07Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif) [http://gcat.davidson.edu/GcatWiki/images/6/6f/3SAT_ppt-1-.ppt]

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T18:06:00Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif) [http://gcat.davidson.edu/GcatWiki/images/6/6f/3SAT ppt-1-.ppt]

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T18:04:37Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif) http://gcat.davidson.edu/GcatWiki/images/6/6f/3SAT ppt-1-.ppt

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T18:00:39Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif)

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

File:File.ogg

2009-05-20T17:59:54Z

Cdavis21:

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T17:59:16Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif)[[Media:file.ogg]]

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T17:57:45Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif)
[[Image:3sat ppt]]

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T17:56:39Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif)
[Image:]

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T17:55:18Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif)
[Image:3SAT ppt-1-.ppt]

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

Can we solve a 3-SAT problem with supressor logic?

2009-05-20T17:53:29Z

Cdavis21: /* What is suppressor logic? */

== What is the 3-SAT problem? ==

''Advantages/Disadvantages to using the 3 SAT problem''

Advantages:

1) Seems to have an equal balance of mathematics and biology.

2) Eventually, there is promise to make the problem difficult enough whereas a computer would have a hard time solving it.

3) Introducing a new concept (supressor supressor logic)

Disadvantages:

1) Using the same concept/idea that has been used by previous Missouri Western/Davidson iGEM teams. (XOR gate)

2) Finding enough for everyone to work on for the entire summer.

3) Finding a promoter that is turned on without the use of a small particle.

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques. 

The 10 clause 3-SAT problem solved in the paper is:

(a or b or –c) 
(a or c or d) 
(a or –c or –d) 
(-a or –c or d) 
(a or –c or e) 
(a or d or –f) 
(-a or c or d) 
(a or c or –d) 
(-a or –c or –d) 
(-a or c or –d) 

note: this problem uses inputs of a, b, c, d, e, -a, -c, -d, -f (f, -b, -e are not used)

== What is suppressor logic? ==

Will's Version of the PPT for SSL 3SAT
[http://gcat.davidson.edu/GcatWiki/images/6/6f/Will%27s_SSL_3SAT.ppt Will's Version of the PPT for SSL 3SAT]

Powerpoint on 3SAT SSL (Bryce and Clif)
[3SAT_ppt-1-.ppt]

'''Suppressor Logic uses suppressor tRNAs as inputs to avoid frameshift mutations in the production of an output amino acid sequence'''

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track. 

If more than one frameshift mutation is introduced into a coding sequence, then logical operators can be encoded. Suppressor a binds to CCCG, supressor b binds to CUGC, and suppressor c binds to ACCG below: 

# AUG CCCG CUG ... rest of Gene 
# AUG CCCG CUGC CUG ... rest of Gene 
# AUG CCCG gg CUGC AGG ... rest of Gene 
# AUG CCCG gg CUGC gg ACCG AGG ... rest of Gene 
 
{| align="center" border="1" cellpadding="8" cellspacing="0" style="background:lightcyan; font-weight:bold; text-align:center; width:45%"
|+
|- style="background:paleturquoise"
! style="width:15%" | Construct
! style="width:15%" | Gene
! style="width:15%" | Logical operation
|-
| 1 || Phenotype || a
|-
| 2 || Phenotype || a AND b
|-
| 3 || Phenotype || a OR b
|-
| 4 || Phenotype || a OR b OR c
|-
| 1 || Repressor|| NOT a
|-
| 2 || Repressor|| a NAND b
|-
| 3 || Repressor|| a NOR b
|-
| 4 || Repressor|| NOT (a OR b OR c)
|}

'''Logical clauses can be connected by AND operators if the proteins produced are part of a biochemical pathway.''' In this case, a AND b AND c: 

<center> [[Image:suppressor2.GIF]] </center>

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

LC using 4-base frameshift mutations: cag CCCG aa GGGC tt GTTG cag (may also have any multiple of 3 bases between the mutations) 

LC using 5-base frameshift mutations: cag CCCGC a GGGCG t GTTGC cag (may also have any multiple of 3 bases between the mutations) 

This makes an XOR logic gate as only one suppressor can be used at one time to maintain the reading frame.

Logical expression (LE) = string of LCs connected by AND 
The design below encodes LC1 AND LC2 AND LC3 

<center> [[Image:logical expression2.GIF]] </center>

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs. Suppressors a and g represent 1 and 0 for the first input; suppressors b and h are 1 and 0 for the second input; etc. up to the sixth input with suppressors f and l (lower case L). The triangles are hix sites for Hin recombination. Whichever of the two suppressor tRNAs in an input pair is facing forward determines whether the value of that input is 1 or 0.

<center> [[Image:suppressor inputs.GIF]] </center>

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1" In order to do this, each of the activators below must turn on a repressor that turns off Hin production. Then if one of the activators is not made, Hin will be made, and new inputs will be established.

<center> [[Image:LE circuit.GIF]] </center>

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

== Things we need to learn about ==

'''Note: We would need 12 different frameshift suppressor tRNAs to encode the Sakamoto 3-SAT problem'''

'''Discovery of frameshift tRNAs. How many are known?''' 

[http://gcat.davidson.edu/GcatWiki/images/5/59/MaglierySchultz2001JMB.pdf Magliery, Anderson, Schultz]

Library approach used to discover efficient suppressors of four-base codons AGGA, UAGA, CCCU, and CUAG using mutated versions of serine tRNA.

[http://gcat.davidson.edu/GcatWiki/images/6/6f/AndersonSchultz2002ChemBiol.pdf Anderson, Maglieri, Schultz]

Signals for translational bypassing (slipping and hopping): mRNA secondary structure, "hungry" (underused) codons, upstream Shine-Dalgarno-like (RBS) sequences 
Library approach extended in order to discover frameshift suppressor tRNAs with anticodons of size two to six bases. Two-base and six-base suppressors were not found, but the following five-base suppressors were found:

<center> [[Image:Table2_Anderson2002.GIF]] </center>

[http://gcat.davidson.edu/GcatWiki/images/a/ac/Dunham_2009_tRNA_structure.pdf Dunham et al.]

[http://gcat.davidson.edu/GcatWiki/images/c/c3/Hohsaka_2009_in_vitro_tln.pdf Hohsaka et al.]

'''Processing of tRNA precursors in E. coli''' 

[http://www.nature.com/embor/journal/v2/n1/full/embor501.html Mörl and Marchfelder] describe processing 

<center> [[Image:the_final_cut.GIF]] </center>

'''How many different frameshift suppressor tRNAs can be in one cell without causing toxicity?'''

Can we have twelve of them? If not, what is the maximum? Is there an impact of codon usage on this?

Can we have both 4- and 5-base suppressor tRNAs in one cell?

Are there particular combinations of suppressors that are more tolerated than others?

 
'''How many frameshift mutations can be suppressed in a single gene?'''

How close together can the frameshift mutations of a given LC be?

Can we mix 4- and 5-base frameshift mutations in a given LC?

File:3SAT ppt-1-.ppt

2009-05-20T17:50:54Z

Cdavis21:

3SAT

2009-05-20T17:49:12Z

Cdavis21:

[[Image:]]

Missouri Western/Davidson iGEM2009

2009-05-14T14:10:55Z

Cdavis21:

This space will be used starting April, 2009 for brainstorming and a shared whiteboard space.

[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/Davidson_Protocols Davidson Lab Protocols] 
[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/MWSU_protocols MWSU Lab Protocols] 
[http://gcat.davidson.edu/sybr-u/bmc.html BioMath Connections Page] 
[http://gcat.davidson.edu/GCATalog-r2.1/GCATalog.htm GCAT-alog Freezer Stocks] 

We need to learn more about these topics:
<center>'''Biology-based'''</center>

#[[How is msDNA normally produced?]] Olivia/Alyndria
#[[How many copies are carried per cell?]] Alyndria
#[[What role can physical modeling of protein structure play in our project?]] Romina
#[[What role can physical modeling of proteins play in our project? Eric Sawyer]]
#[[What other cool reporters are there? (Discrete On/Off or Continuous) Bryce Szczepanik]]
#[[Can we use promoter strength/opposite directions to subtract? Clif Davis]]
#[[What is msDNA?]]
#[[Can we use suppressor tRNAs to encode logical operators (suppressor suppressor logic, SSL)?]]
#[[How is msDNA stored in E. coli?]] Olivia
#[[What is the sequence of bacterial reverse transcriptase and can we clone that gene?]] Shamita
#[[Can we redesign the normal msDNA pathway to produce new segments of DNA of our choosing?]] All
#[[What are other available reverse transcriptases?]] Leland
#[[What other math problems (e.g. NP- complete) are accessible to us? Siya Sun]]
#[[What is the relationship between 3-SAT and map coloring? Ashley Schnoor]]
#[[What activators are there that turn on a promoter without any help?]]
#[[Can we use protein interactions to compute? (Post-translation, proteases, quaternary structure) Will Vernon]]
#[[Could we do something with clocks/counting?]]
#[[Could we have/use multiple synthetic organelles in a cell?]]
#[[What ideas from previous iGEM teams are useful to us?]]

<center>'''Math-based'''</center>
#[[Can we get bacteria to solve a problem large enough to challenge a person?]]
#What interesting challenges or problems does origami offer?
#Can we produce a series of increasingly difficult goals that might be possible to produce in the lab?
#What has been done before and how can we improve upon that?
#We can perform some pilot experiments using synthesized DNA and later switch to msDNA (maybe).
#Can we address the Boolean Satisfiability (SAT) problem with a bacterial computer?
#How has 3SAT been addressed with a DNA computer? Can we use those methods?

#Can we get bacteria to solve a problem large enough to challenge a computer (probably not, but it is fun to think about)?
#What are some linear algebra applications for DNA origami?
#How can we use origami to solve 3-SAT problems?

<center>'''Behavior-based'''</center>
#[[What constructs are we testing?]]
#[[What school districts do we have access to?]]
#[[Where is the Synthetic Biology page we want high school teachers to use after the survey?]]
#[[Do you need any more input from the veterans before the survey is ready?]]

<center>'''Solving The 3SAT Problem Using Suppressor Logic'''</center>
#[[Can we solve a 3-SAT problem with supressor logic?]]

Missouri Western/Davidson iGEM2009

2009-05-14T13:58:46Z

Cdavis21:

This space will be used starting April, 2009 for brainstorming and a shared whiteboard space.

[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/Davidson_Protocols Davidson Lab Protocols] 
[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/MWSU_protocols MWSU Lab Protocols] 
[http://gcat.davidson.edu/sybr-u/bmc.html BioMath Connections Page] 
[http://gcat.davidson.edu/GCATalog-r2.1/GCATalog.htm GCAT-alog Freezer Stocks] 

We need to learn more about these topics:
<center>'''Biology-based'''</center>

#[[How is msDNA normally produced?]] Olivia/Alyndria
#[[How many copies are carried per cell?]] Alyndria
#[[What role can physical modeling of protein structure play in our project?]] Romina
#[[What role can physical modeling of proteins play in our project? Eric Sawyer]]
#[[What other cool reporters are there? (Discrete On/Off or Continuous) Bryce Szczepanik]]
#[[Can we use promoter strength/opposite directions to subtract? Clif Davis]]
#[[What is msDNA?]]
#[[Can we use suppressor tRNAs to encode logical operators (suppressor suppressor logic, SSL)?]]
#[[How is msDNA stored in E. coli?]] Olivia
#[[What is the sequence of bacterial reverse transcriptase and can we clone that gene?]] Shamita
#[[Can we redesign the normal msDNA pathway to produce new segments of DNA of our choosing?]] All
#[[What are other available reverse transcriptases?]] Leland
#[[What other math problems (e.g. NP- complete) are accessible to us? Siya Sun]]
#[[What is the relationship between 3-SAT and map coloring? Ashley Schnoor]]
#[[What activators are there that turn on a promoter without any help?]]
#[[Can we use protein interactions to compute? (Post-translation, proteases, quaternary structure) Will Vernon]]
#[[Could we do something with clocks/counting?]]
#[[Could we have/use multiple synthetic organelles in a cell?]]
#[[What ideas from previous iGEM teams are useful to us?]]

<center>'''Math-based'''</center>
#What interesting challenges or problems does origami offer?
#Can we produce a series of increasingly difficult goals that might be possible to produce in the lab?
#What has been done before and how can we improve upon that?
#We can perform some pilot experiments using synthesized DNA and later switch to msDNA (maybe).
#Can we address the Boolean Satisfiability (SAT) problem with a bacterial computer?
#How has 3SAT been addressed with a DNA computer? Can we use those methods?
#[[Can we get bacteria to solve a problem large enough to challenge a person?]]
#Can we get bacteria to solve a problem large enough to challenge a computer (probably not, but it is fun to think about)?
#What are some linear algebra applications for DNA origami?
#How can we use origami to solve 3-SAT problems?

<center>'''Behavior-based'''</center>
#[[What constructs are we testing?]]
#[[What school districts do we have access to?]]
#[[Where is the Synthetic Biology page we want high school teachers to use after the survey?]]
#[[Do you need any more input from the veterans before the survey is ready?]]

<center>'''Solving The 3SAT Problem Using Suppressor Logic'''</center>
#[[Can we solve a 3-SAT problem with supressor logic?]]

Missouri Western/Davidson iGEM2009

2009-05-14T13:58:08Z

Cdavis21:

This space will be used starting April, 2009 for brainstorming and a shared whiteboard space.

[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/Davidson_Protocols Davidson Lab Protocols] 
[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/MWSU_protocols MWSU Lab Protocols] 
[http://gcat.davidson.edu/sybr-u/bmc.html BioMath Connections Page] 
[http://gcat.davidson.edu/GCATalog-r2.1/GCATalog.htm GCAT-alog Freezer Stocks] 

We need to learn more about these topics:
<center>'''Biology-based'''</center>

#[[How is msDNA normally produced?]] Olivia/Alyndria
#[[How many copies are carried per cell?]] Alyndria
#[[What role can physical modeling of protein structure play in our project?]] Romina
#[[What role can physical modeling of proteins play in our project? Eric Sawyer]]
#[[What other cool reporters are there? (Discrete On/Off or Continuous) Bryce Szczepanik]]
#[[Can we use promoter strength/opposite directions to subtract? Clif Davis]]
#[[What is msDNA?]]
#[[Can we use suppressor tRNAs to encode logical operators (suppressor suppressor logic, SSL)?]]
#[[How is msDNA stored in E. coli?]] Olivia
#[[What is the sequence of bacterial reverse transcriptase and can we clone that gene?]] Shamita
#[[Can we redesign the normal msDNA pathway to produce new segments of DNA of our choosing?]] All
#[[What are other available reverse transcriptases?]] Leland
#[[Can we solve a 3-SAT problem with supressor logic?]]
#[[What other math problems (e.g. NP- complete) are accessible to us? Siya Sun]]
#[[What is the relationship between 3-SAT and map coloring? Ashley Schnoor]]
#[[What activators are there that turn on a promoter without any help?]]
#[[Can we use protein interactions to compute? (Post-translation, proteases, quaternary structure) Will Vernon]]
#[[Could we do something with clocks/counting?]]
#[[Could we have/use multiple synthetic organelles in a cell?]]
#[[What ideas from previous iGEM teams are useful to us?]]

<center>'''Math-based'''</center>
#What interesting challenges or problems does origami offer?
#Can we produce a series of increasingly difficult goals that might be possible to produce in the lab?
#What has been done before and how can we improve upon that?
#We can perform some pilot experiments using synthesized DNA and later switch to msDNA (maybe).
#Can we address the Boolean Satisfiability (SAT) problem with a bacterial computer?
#How has 3SAT been addressed with a DNA computer? Can we use those methods?
#[[Can we get bacteria to solve a problem large enough to challenge a person?]]
#Can we get bacteria to solve a problem large enough to challenge a computer (probably not, but it is fun to think about)?
#What are some linear algebra applications for DNA origami?
#How can we use origami to solve 3-SAT problems?

<center>'''Behavior-based'''</center>
#[[What constructs are we testing?]]
#[[What school districts do we have access to?]]
#[[Where is the Synthetic Biology page we want high school teachers to use after the survey?]]
#[[Do you need any more input from the veterans before the survey is ready?]]

<center>'''Solving The 3SAT Problem using Suppressor Logic'''</center>

Missouri Western/Davidson iGEM2009

2009-05-14T13:55:33Z

Cdavis21:

This space will be used starting April, 2009 for brainstorming and a shared whiteboard space.

[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/Davidson_Protocols Davidson Lab Protocols] 
[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/MWSU_protocols MWSU Lab Protocols] 
[http://gcat.davidson.edu/sybr-u/bmc.html BioMath Connections Page] 
[http://gcat.davidson.edu/GCATalog-r2.1/GCATalog.htm GCAT-alog Freezer Stocks] 

We need to learn more about these topics:
<center>'''Biology-based'''</center>

#[[How is msDNA normally produced?]] Olivia/Alyndria
#[[How many copies are carried per cell?]] Alyndria
#[[What role can physical modeling of protein structure play in our project?]] Romina
#[[What role can physical modeling of proteins play in our project? Eric Sawyer]]
#[[What other cool reporters are there? (Discrete On/Off or Continuous) Bryce Szczepanik]]
#[[Can we use promoter strength/opposite directions to subtract? Clif Davis]]
#[[What is msDNA?]]
#[[Can we use suppressor tRNAs to encode logical operators (suppressor suppressor logic, SSL)?]]
#[[How is msDNA stored in E. coli?]] Olivia
#[[What is the sequence of bacterial reverse transcriptase and can we clone that gene?]] Shamita
#[[Can we redesign the normal msDNA pathway to produce new segments of DNA of our choosing?]] All
#[[What are other available reverse transcriptases?]] Leland
#[[Can we solve a 3-SAT problem with supressor logic?]]
#[[What other math problems (e.g. NP- complete) are accessible to us? Siya Sun]]
#[[What is the relationship between 3-SAT and map coloring? Ashley Schnoor]]
#[[What activators are there that turn on a promoter without any help?]]
#[[Can we use protein interactions to compute? (Post-translation, proteases, quaternary structure) Will Vernon]]
#[[Could we do something with clocks/counting?]]
#[[Could we have/use multiple synthetic organelles in a cell?]]
#[[What ideas from previous iGEM teams are useful to us?]]

<center>'''Math-based'''</center>
#What interesting challenges or problems does origami offer?
#Can we produce a series of increasingly difficult goals that might be possible to produce in the lab?
#What has been done before and how can we improve upon that?
#We can perform some pilot experiments using synthesized DNA and later switch to msDNA (maybe).
#Can we address the Boolean Satisfiability (SAT) problem with a bacterial computer?
#How has 3SAT been addressed with a DNA computer? Can we use those methods?
#[[Can we get bacteria to solve a problem large enough to challenge a person?]]
#Can we get bacteria to solve a problem large enough to challenge a computer (probably not, but it is fun to think about)?
#What are some linear algebra applications for DNA origami?
#How can we use origami to solve 3-SAT problems?

<center>'''Behavior-based'''</center>
#[[What constructs are we testing?]]
#[[What school districts do we have access to?]]
#[[Where is the Synthetic Biology page we want high school teachers to use after the survey?]]
#[[Do you need any more input from the veterans before the survey is ready?]]

Missouri Western/Davidson iGEM2009

2009-05-11T20:51:10Z

Cdavis21:

This space will be used starting April, 2009 for brainstorming and a shared whiteboard space.

[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/Davidson_Protocols Davidson Lab Protocols] 
[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/MWSU_protocols MWSU Lab Protocols] 
[http://gcat.davidson.edu/sybr-u/bmc.html BioMath Connections Page] 
[http://gcat.davidson.edu/GCATalog-r2.1/GCATalog.htm GCAT-along Freezer Stocks] 

We need to learn more about these topics:
<center>'''Biology-based'''</center>
#[[What is msDNA?]]
#[[How is msDNA normally produced?]] Olivia/Alyndria
#[[How is msDNA stored in E. coli?]] Olivia
#[[How many copies are carried per cell?]] Alyndria
#[[What is the sequence of bacterial reverse transcriptase and can we clone that gene?]] Shamita
#[[Can we redesign the normal msDNA pathway to produce new segments of DNA of our choosing?]] All
#[[Can we use suppressor tRNAs to encode logical operators (suppressor suppressor logic, SSL)?]]
#[[What are other available reverse transcriptases?]] Leland
#[[Can we solve a 3-SAT problem with supressor logic?]]
#[[What role can physical modeling of protein structure play in our project?]] Romina
#[[What other math problems (e.g. NP- complete) are accessible to us? Siya Sun]]
#[[What is the relationship between 3-SAT and map coloring? Ashley Schnoor]]
#[[What role can physical modeling of proteins play in our project? Eric Sawyer]]
#[[What activators are there that turn on a promoter without any help?]]
#[[What other cool reporters are there? (Discrete On/Off or Continuous) Bryce Szczepanik]]
#[[Can we use promoter strength/opposite directions to subtract? Clif Davis]]
#[[Can we use protein interactions to compute? (Post-translation, proteases, quaternary structure) Will Vernon]]
#[[Could we do something with clocks/counting?]]
#[[Could we have/use multiple synthetic organelles in a cell?]]
#[[What ideas from previous iGEM teams are useful to us?]]

<center>'''Math-based'''</center>
#What interesting challenges or problems does origami offer?
#Can we produce a series of increasingly difficult goals that might be possible to produce in the lab?
#What has been done before and how can we improve upon that?
#We can perform some pilot experiments using synthesized DNA and later switch to msDNA (maybe).
#Can we address the Boolean Satisfiability (SAT) problem with a bacterial computer?
#How has 3SAT been addressed with a DNA computer? Can we use those methods?
#[[Can we get bacteria to solve a problem large enough to challenge a person?]]
#Can we get bacteria to solve a problem large enough to challenge a computer (probably not, but it is fun to think about)?
#What are some linear algebra applications for DNA origami?
#How can we use origami to solve 3-SAT problems?

<center>'''Behavior-based'''</center>
#[[What constructs are we testing?]]
#[[What school districts do we have access to?]]
#[[Where is the Synthetic Biology page we want high school teachers to use after the survey?]]
#[[Do you need any more input from the veterans before the survey is ready?]]

Missouri Western/Davidson iGEM2009

2009-05-11T20:49:15Z

Cdavis21:

This space will be used starting April, 2009 for brainstorming and a shared whiteboard space.

[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/Davidson_Protocols Davidson Lab Protocols] 
[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/MWSU_protocols MWSU Lab Protocols] 
[http://gcat.davidson.edu/sybr-u/bmc.html BioMath Connections Page] 
[http://gcat.davidson.edu/GCATalog-r2.1/GCATalog.htm GCAT-along Freezer Stocks] 

We need to learn more about these topics:
<center>'''Biology-based'''</center>
#[[What is msDNA?]]
#[[How is msDNA normally produced?]] Olivia/Alyndria
#[[How is msDNA stored in E. coli?]] Olivia
#[[How many copies are carried per cell?]] Alyndria
#[[What is the sequence of bacterial reverse transcriptase and can we clone that gene?]] Shamita
#[[Can we redesign the normal msDNA pathway to produce new segments of DNA of our choosing?]] All
#[[Can we use suppressor tRNAs to encode logical operators (suppressor suppressor logic, SSL)?]]
#[[What are other available reverse transcriptases?]] Leland
#[[Can we solve a 3-SAT problem with supressor logic?]]
#[[What role can physical modeling of protein structure play in our project?]] Romina
#[[Could we have/use multiple synthetic organelles in a cell?]]
#[[What ideas from previous iGEM teams are useful to us?]]
#[[What other math problems (e.g. NP- complete) are accessible to us? Siya Sun]]
#[[What is the relationship between 3-SAT and map coloring? Ashley Schnoor]]
#[[Could we do something with clocks/counting?]]
#[[What role can physical modeling of proteins play in our project? Eric Sawyer]]
#[[What activators are there that turn on a promoter without any help?]]
#[[What other cool reporters are there? (Discrete On/Off or Continuous) Bryce Szczepanik]]
#[[Can we use promoter strength/opposite directions to subtract? Clif Davis]]
#[[Can we use protein interactions to compute? (Post-translation, proteases, quaternary structure) Will Vernon]]

<center>'''Math-based'''</center>
#What interesting challenges or problems does origami offer?
#Can we produce a series of increasingly difficult goals that might be possible to produce in the lab?
#What has been done before and how can we improve upon that?
#We can perform some pilot experiments using synthesized DNA and later switch to msDNA (maybe).
#Can we address the Boolean Satisfiability (SAT) problem with a bacterial computer?
#How has 3SAT been addressed with a DNA computer? Can we use those methods?
#[[Can we get bacteria to solve a problem large enough to challenge a person?]]
#Can we get bacteria to solve a problem large enough to challenge a computer (probably not, but it is fun to think about)?
#What are some linear algebra applications for DNA origami?
#How can we use origami to solve 3-SAT problems?

<center>'''Behavior-based'''</center>
#[[What constructs are we testing?]]
#[[What school districts do we have access to?]]
#[[Where is the Synthetic Biology page we want high school teachers to use after the survey?]]
#[[Do you need any more input from the veterans before the survey is ready?]]

Missouri Western/Davidson iGEM2009

2009-05-11T20:47:06Z

Cdavis21:

This space will be used starting April, 2009 for brainstorming and a shared whiteboard space.

[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/Davidson_Protocols Davidson Lab Protocols] 
[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/MWSU_protocols MWSU Lab Protocols] 
[http://gcat.davidson.edu/sybr-u/bmc.html BioMath Connections Page] 
[http://gcat.davidson.edu/GCATalog-r2.1/GCATalog.htm GCAT-along Freezer Stocks] 

We need to learn more about these topics:
<center>'''Biology-based'''</center>
#[[What is msDNA?]]
#[[How is msDNA normally produced?]] Olivia/Alyndria
#[[How is msDNA stored in E. coli?]] Olivia
#[[How many copies are carried per cell?]] Alyndria
#[[What is the sequence of bacterial reverse transcriptase and can we clone that gene?]] Shamita
#[[Can we redesign the normal msDNA pathway to produce new segments of DNA of our choosing?]] All
#[[Can we use suppressor tRNAs to encode logical operators (suppressor suppressor logic, SSL)?]]
#[[What are other available reverse transcriptases?]] Leland
#[[Can we solve a 3-SAT problem with supressor logic?]]
#[[What role can physical modeling of protein structure play in our project?]] Romina
#[[Could we have/use multiple synthetic organelles in a cell?]]
#[[What ideas from previous iGEM teams are useful to us?]]
#[[What other math problems (e.g. NP- complete) are accessible to us? Siya Sun]]
#[[What is the relationship between 3-SAT and map coloring? Ashley Schnoor]]
#[[Could we do something with clocks/counting?]]
#[[What role can physical modeling of proteins play in our project? Eric Sawyer]]
#[[What activators are there that turn on a promoter without any help?]]
#[[What other cool reporters are there? (Discrete On/Off or Continuous) Bryce Szczepanik]]
#[[Can we use promoter strength/opposite directions to subtract? Clif Davis]]
#[[Can we use protein interactions to compute? (Post-translation, proteases, quaternary structure) Will Vernon]]

<center>'''Math-based'''</center>
#What interesting challenges or problems does origami offer?
#Can we produce a series of increasingly difficult goals that might be possible to produce in the lab?
#What has been done before and how can we improve upon that?
#We can perform some pilot experiments using synthesized DNA and later switch to msDNA (maybe).
#Can we address the Boolean Satisfiability (SAT) problem with a bacterial computer?
#How has 3SAT been addressed with a DNA computer? Can we use those methods?
#[[Can we get bacteria to solve a problem large enough to challenge a person?]]
#Can we get bacteria to solve a problem large enough to challenge a computer (probably not, but it is fun to think about)?
#What are some linear algebra applications for DNA origami?
#How can we use origami to solve 3-SAT problems?

<center>'''Behavior-based'''</center>
#[[What constructs are we testing?]]
#[[What school districts do we have access to?]]
#[[Where is the Synthetic Biology page we want high school teachers to use after the survey?]]
#[[Do you need any more input from the veterans before the survey is ready?]]

<center>'''Questions from BMC 4/17/09'''</center>

(In no particular order)

#[[Could we have/use multiple synthetic organelles in a cell?]]
#[[What ideas from previous iGEM teams are useful to us?]]
#[[What other math problems (e.g. NP- complete) are accessible to us? Siya Sun]]
#[[What is the relationship between 3-SAT and map coloring? Ashley Schnoor]]
#[[Could we do something with clocks/counting?]]
#[[What role can physical modeling of proteins play in our project? Eric Sawyer]]
#[[What activators are there that turn on a promoter without any help?]]
#[[What other cool reporters are there? (Discrete On/Off or Continuous) Bryce Szczepanik]]
#[[Can we use promoter strength/opposite directions to subtract? Clif Davis]]
#[[Can we use protein interactions to compute? (Post-translation, proteases, quaternary structure) Will Vernon]]

Missouri Western/Davidson iGEM2009

2009-05-11T20:46:10Z

Cdavis21:

This space will be used starting April, 2009 for brainstorming and a shared whiteboard space.

[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/Davidson_Protocols Davidson Lab Protocols] 
[http://gcat.davidson.edu/GcatWiki/index.php/Davidson_Missouri_W/MWSU_protocols MWSU Lab Protocols] 
[http://gcat.davidson.edu/sybr-u/bmc.html BioMath Connections Page] 
[http://gcat.davidson.edu/GCATalog-r2.1/GCATalog.htm GCAT-along Freezer Stocks] 

We need to learn more about these topics:
<center>'''Biology-based'''</center>
#[[What is msDNA?]]
#[[How is msDNA normally produced?]] Olivia/Alyndria
#[[How is msDNA stored in E. coli?]] Olivia
#[[How many copies are carried per cell?]] Alyndria
#[[What is the sequence of bacterial reverse transcriptase and can we clone that gene?]] Shamita
#[[Can we redesign the normal msDNA pathway to produce new segments of DNA of our choosing?]] All
#[[Can we use suppressor tRNAs to encode logical operators (suppressor suppressor logic, SSL)?]]
#[[What are other available reverse transcriptases?]] Leland
#[[Can we solve a 3-SAT problem with supressor logic?]]
#[[What role can physical modeling of protein structure play in our project?]] Romina
#[[Can we use promoter strength/opposite directions to subtract? Clif Davis]]

<center>'''Math-based'''</center>
#What interesting challenges or problems does origami offer?
#Can we produce a series of increasingly difficult goals that might be possible to produce in the lab?
#What has been done before and how can we improve upon that?
#We can perform some pilot experiments using synthesized DNA and later switch to msDNA (maybe).
#Can we address the Boolean Satisfiability (SAT) problem with a bacterial computer?
#How has 3SAT been addressed with a DNA computer? Can we use those methods?
#[[Can we get bacteria to solve a problem large enough to challenge a person?]]
#Can we get bacteria to solve a problem large enough to challenge a computer (probably not, but it is fun to think about)?
#What are some linear algebra applications for DNA origami?
#How can we use origami to solve 3-SAT problems?

<center>'''Behavior-based'''</center>
#[[What constructs are we testing?]]
#[[What school districts do we have access to?]]
#[[Where is the Synthetic Biology page we want high school teachers to use after the survey?]]
#[[Do you need any more input from the veterans before the survey is ready?]]

<center>'''Questions from BMC 4/17/09'''</center>

(In no particular order)

#[[Could we have/use multiple synthetic organelles in a cell?]]
#[[What ideas from previous iGEM teams are useful to us?]]
#[[What other math problems (e.g. NP- complete) are accessible to us? Siya Sun]]
#[[What is the relationship between 3-SAT and map coloring? Ashley Schnoor]]
#[[Could we do something with clocks/counting?]]
#[[What role can physical modeling of proteins play in our project? Eric Sawyer]]
#[[What activators are there that turn on a promoter without any help?]]
#[[What other cool reporters are there? (Discrete On/Off or Continuous) Bryce Szczepanik]]
#[[Can we use promoter strength/opposite directions to subtract? Clif Davis]]
#[[Can we use protein interactions to compute? (Post-translation, proteases, quaternary structure) Will Vernon]]

File:Color2.GIF

2009-05-11T16:02:58Z

Cdavis21:

Can we use promoter strength/opposite directions to subtract? Clif Davis

2009-05-11T16:02:48Z

Cdavis21: /* Subtraction Using Promoter Strength */

=== Promoter Strength ===

I found some valuable information on the Cornell University Website listed below.
“The degree to which a given promoter conforms to the consensus sequence determines the strength of that promoter. The closer the sequence to the consensus, the stronger the promoter will be and the more frequently transcription will occur at that promoter.”
“Promoter strength is important because it determines how often a given mRNA sequence is transcribed, effectively giving higher priority for transcription to some genes over others. A gene that codes for a protein that is required in large quantities, for example, might be expected to have a relatively strong promoter.”

=== References ===

http://www.biog1105-1106.org/demos/106/unit02/3c.promoterstrength.html

=== Subtraction Using Promoter Strength ===

Now say we experiment with some different strength promoters, and use them with the manufacturing of some fluorescent protein. We could then grade them by measuring the color formation against each other. Say we had Yellow Fluorescent Protein (YFP) and Green Fluorescent Protein (GFP) and used our different promoters to make some of each. If we use two different promoters, and the end result is a greenyellow color, then it would be safe to assume that the promoter on the GFP gene is stronger than the other promoter. Maybe if we got it just right, with equal promoters on each gene, we could come up with a blue color on our plates. I included some sample colors to give more of a visual effect.

<center> [[Image:color2.GIF]] </center>

Obviously there are too many different degrees of color for people to distinguish. We may run into a problem in there being enough difference between the two promoters to tell the difference in color.

Once we grade the promoters we are using, maybe on a 1-10 scale. We should be able to make a key of some kind. A difference of 1 should always look like this, or a difference of 7 should look like this. We could start there with our math problems and see what happens. The more promoters we can use the bigger the difference can be in strengths.

Can we use promoter strength/opposite directions to subtract? Clif Davis

2009-05-11T15:59:14Z

Cdavis21: /* Subtraction Using Promoter Strength */

=== Promoter Strength ===

I found some valuable information on the Cornell University Website listed below.
“The degree to which a given promoter conforms to the consensus sequence determines the strength of that promoter. The closer the sequence to the consensus, the stronger the promoter will be and the more frequently transcription will occur at that promoter.”
“Promoter strength is important because it determines how often a given mRNA sequence is transcribed, effectively giving higher priority for transcription to some genes over others. A gene that codes for a protein that is required in large quantities, for example, might be expected to have a relatively strong promoter.”

=== References ===

http://www.biog1105-1106.org/demos/106/unit02/3c.promoterstrength.html

=== Subtraction Using Promoter Strength ===

Now say we experiment with some different strength promoters, and use them with the manufacturing of some fluorescent protein. We could then grade them by measuring the color formation against each other. Say we had Yellow Fluorescent Protein (YFP) and Green Fluorescent Protein (GFP) and used our different promoters to make some of each. If we use two different promoters, and the end result is a greenyellow color, then it would be safe to assume that the promoter on the GFP gene is stronger than the other promoter. Maybe if we got it just right, with equal promoters on each gene, we could come up with a blue color on our plates. I included some sample colors to give more of a visual effect.

<center> [[Image:color.GIF]] </center>

Obviously there are too many different degrees of color for people to distinguish. We may run into a problem in there being enough difference between the two promoters to tell the difference in color.

Once we grade the promoters we are using, maybe on a 1-10 scale. We should be able to make a key of some kind. A difference of 1 should always look like this, or a difference of 7 should look like this. We could start there with our math problems and see what happens. The more promoters we can use the bigger the difference can be in strengths.

File:Color.GIF

2009-05-11T15:48:25Z

Cdavis21:

Can we use promoter strength/opposite directions to subtract? Clif Davis

2009-05-11T15:48:09Z

Cdavis21: /* Subtraction Using Promoter Strength */

Can we use promoter strength/opposite directions to subtract? Clif Davis

2009-05-11T15:45:21Z

Cdavis21: /* Promoter Strength */

Can we use promoter strength/opposite directions to subtract? Clif Davis

2009-05-11T15:43:53Z

Cdavis21:

=== Promoter Strength ===

I found some valuable information on the Cornell University Website listed below.
“The degree to which a given promoter conforms to the consensus sequence determines the strength of that promoter. The closer the sequence to the consensus, the stronger the promoter will be and the more frequently transcription will occur at that promoter.”
“Promoter strength is important because it determines how often a given mRNA sequence is transcribed, effectively giving higher priority for transcription to some genes over others. A gene that codes for a protein that is required in large quantities, for example, might be expected to have a relatively strong promoter.”
http://www.biog1105-1106.org/demos/106/unit02/3c.promoterstrength.html

=== Subtraction Using Promoter Strength ===

Now say we experiment with some different strength promoters, and use them with the manufacturing of some fluorescent protein. We could then grade them by measuring the color formation against each other. Say we had Yellow Fluorescent Protein (YFP) and Green Fluorescent Protein (GFP) and used our different promoters to make some of each. If we use two different promoters, and the end result is a greenyellow color, then it would be safe to assume that the promoter on the GFP gene is stronger than the other promoter. Maybe if we got it just right, with equal promoters on each gene, we could come up with a blue color on our plates. I included some sample colors to give more of a visual effect.

File:Hairpin2.GIF

2009-04-07T20:35:49Z

Cdavis21:

Can we solve a 3-SAT problem with supressor logic?

2009-04-07T20:35:38Z

Cdavis21: /* How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? */

== What is the 3-SAT problem? ==

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin2.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques.

== What is suppressor logic? ==

<center> [[Image:suppressor2.GIF]] </center>

Suppressor Suppressor logic uses suppressor tRNAs to avoid frameshift mutations in an amino acid sequence.

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track.

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

Logical expression (LE) = string of LCs connected by AND

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs.

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1"

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

Can we solve a 3-SAT problem with supressor logic?

2009-04-07T20:35:09Z

Cdavis21: /* How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? */

== What is the 3-SAT problem? ==

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques.

== What is suppressor logic? ==

<center> [[Image:suppressor2.GIF]] </center>

Suppressor Suppressor logic uses suppressor tRNAs to avoid frameshift mutations in an amino acid sequence.

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track.

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

Logical expression (LE) = string of LCs connected by AND

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs.

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1"

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

Can we solve a 3-SAT problem with supressor logic?

2009-04-07T20:34:58Z

Cdavis21: /* How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? */

== What is the 3-SAT problem? ==

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques.

== What is suppressor logic? ==

<center> [[Image:suppressor2.GIF]] </center>

Suppressor Suppressor logic uses suppressor tRNAs to avoid frameshift mutations in an amino acid sequence.

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track.

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

Logical expression (LE) = string of LCs connected by AND

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs.

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1"

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

Can we solve a 3-SAT problem with supressor logic?

2009-04-07T20:33:45Z

Cdavis21: /* How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? */

== What is the 3-SAT problem? ==

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin.GIF]] </center>
Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques.

== What is suppressor logic? ==

<center> [[Image:suppressor2.GIF]] </center>

Suppressor Suppressor logic uses suppressor tRNAs to avoid frameshift mutations in an amino acid sequence.

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track.

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

Logical expression (LE) = string of LCs connected by AND

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs.

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1"

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

Can we solve a 3-SAT problem with supressor logic?

2009-04-07T20:33:16Z

Cdavis21: /* How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? */

== What is the 3-SAT problem? ==

== How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? ==

[http://gcat.davidson.edu/GcatWiki/images/3/39/Sakamoto_DNA_computer_3SAT.pdf Sakamoto et al.]

<center> [[Image:hairpin.GIF]] </center>

Sakamoto used hairpin formations in single stranded DNA (ssDNA) as a molecular computer. Hairpin structures are formed when complimentary bases on the same strand attach to each other forming a loop. As shown in Picture B above, C and -C are compliments of each other, and bind together. Picture A is what a normal ssDNA should look like. Sakamoto used this self assembly of secondary structures on a satisfiability (SAT) problem. The problem had six inputs and ten clauses. An example of a clause would be (a or b or -c), and could include any combinations of inputs from a to f including -a to -f. If the problem is satisfied, the ssDNA stays in it's normal form. If the problem is not satisfied, the ssDNA forms a hairpin. Later in the paper it explains that the hairpin forming molecules can be removed from the others with certain techniques.

== What is suppressor logic? ==

<center> [[Image:suppressor2.GIF]] </center>

Suppressor Suppressor logic uses suppressor tRNAs to avoid frameshift mutations in an amino acid sequence.

A frameshift is a genetic mutation caused by the addition or deletion of nucelotides to a given sequence which codes for a protein. Since codons are read in a series of three, the addition or deletion of nucleotides will disrupt the reading frame. This disruption will most likely cause the production of a nonfunctional protein.

<center> [[Image:suppressor4.GIF]] </center>

A frameshift occurs and, in this case, a guanine is added to the sequence. If nothing is done, enzyme A will not be made, meaning the clause will not be satisfied.

The suppressor tRNA allows the 4 letter sequence to be read as a single codon, therefore, keeping the protein on track.

== How could suppressor logic be used to solve the Sakamoto 3-SAT problem? ==

'''Definitions'''

Inputs = framshift suppressor tRNAs

Input value = supp a is 1, supp g is 0; supp b is 1, supp h is 0, etc. up to tth 6th pair of f and l

Logical clause (LC) = three inputs connected by OR, eg. (a OR b OR e)

Logical expression (LE) = string of LCs connected by AND

'''Subroutine'''

1. Individual bacteral cells use Hin/hix system to randomly choose of of the 64 possible combinations of 6 inputs.

2. Each bacterial cell carries out the following subroutine on each LC: IF LC=TRUE THEN "check the next LC" ELSEIF LC=FALSE "go get a new set of inputs with step 1"

3. If/when a bacterial cell finds a set of inputs that satisfies the entire LE (ie. a solution to the 3-SAT problem), it will glow green.

File:Hairpin.GIF

2009-04-07T19:56:26Z

Cdavis21:

Can we solve a 3-SAT problem with supressor logic?

2009-04-07T19:55:37Z

Cdavis21: /* How did Sakamoto et al. use a DNA computer to solve a 3-SAT problem? */