== Suppressor Logic Toolkit and Quasi-Hash Function ==

The following slides describe the basic toolkit design and a specific experiment that would result in a quasi-hash function

== Basic Toolkit Design ==

The AND gate was designed by Anderson using termination suppressors, while the new design would use frame shift suppressors.

<center> [[Image:Picture13.gif]] </center>
<center> [[Image:Picture14.gif]] </center>
<center> [[Image:Picture15.gif]] </center>
<center> [[Image:Picture16.gif]] </center>

== Toolkit with Specific Inputs and Outputs ==

PprpB/propionate and luxO/pluxT or pqrr4 are new communication players that would need to be looked into further to verify their usefulness.

See references at bottom of page.

<center> [[Image:andgate.gif]] </center>
<center> [[Image:Picture29.gif]] </center>
<center> [[Image:Picture30.gif]] </center>
<center> [[Image:Picture31.gif]] </center>

== Experimental Design ==

<center> [[Image:Picture21.gif]] </center>
<center> [[Image:Picture22.gif]] </center>

== 434 repressor ==
''Design:''

1. OR1 is needed. the role of OR1 is to help repressor bind to OR2 and to repress the PR;

2. OR2 is needed. The binding of the '''amino-terminal domain''' of 434 repressor to OR2 is necessary and sufficient for activation of PRM transcription;[http://www.pnas.org/content/83/24/9353.full.pdf+html]

3. Need to decrease the binding affinity of OR3. binding to OR3 will cause repression of PRM transcription.Introduce a mutation in OR3 that decreases its affinity for the repressor without disturbing PRM;

4. Changing the sequence of the -35 region of PRM toward consensus greatly increases the ability of RNA polymerase to transcribe this DNA in the absence of the repressor...the mutation decreases the ability of RNA polymerase to initiate transcription at PR [http://www.jbc.org/cgi/content/full/273/37/24165]

http://gcat.davidson.edu/GcatWiki/images/c/ce/434r.JPG

== Parts Needed ==

'''Parts from Registry'''

'''Promoters'''

pR [http://partsregistry.org/wiki/index.php?title=Part:BBa_R0051 R0051]2009 Kit Plate 1 6K pSB1A2

pLacIQ1 [http://partsregistry.org/wiki/index.php/Part:BBa_K091112 K091112]MWSU

pTetR [http://partsregistry.org/wiki/index.php/Part:BBa_R0040 R0040]2009 Kit Plate 1 6I pSB1A2

pLuxR [http://partsregistry.org/wiki/index.php/Part:BBa_R0062 R0062]2009 Kit Plate 1 6O pSB1A2

pMnt [http://partsregistry.org/wiki/index.php?title=Part:BBa_R0073 R0073]2009 Kit Plate 1 16B pSB2K3

'''Genes'''

RBS+lamba cI+TT [http://partsregistry.org/wiki/index.php/Part:BBa_P0451 P0451]2009 Kit Plate 2 1N pSB1AK3

pBAD+RBS+LacI+TT [http://partsregistry.org/wiki/index.php/Part:BBa_S03980 S03980]MWSU

pBAD+RBS+tetR+TT [http://partsregistry.org/wiki/index.php/Part:BBa_K091145 K091145]MWSU

pBAD+RBS+luxR+TT [http://partsregistry.org/wiki/index.php/Part:BBa_K091179 K091179]MWSU

RBS(B0030)+GFP [http://partsregistry.org/wiki/index.php/Part:BBa_E5500 E5500]2007 Parts Kit Plate 1 9J pSB1A2

RBS(B0034)+RFP MWSU

Mnt [http://partsregistry.org/wiki/index.php?title=Part:BBa_C0072 C0072]2009 Kit Plate 1 12L pSB2K3

LuxO-P [http://partsregistry.org/wiki/index.php?title=Part:BBa_K131022 K131022]2009 Kit Plate 3 8K pSB1AC3

pqrr4 [http://partsregistry.org/Part:BBa_K131017 K131017]

'''Parts that need modified'''

LuxO-P [http://partsregistry.org/wiki/index.php?title=Part:BBa_K131022 K131022]2009 Kit Plate 3 8K pSB1AC3

RBS+lamba cI+TT [http://partsregistry.org/wiki/index.php/Part:BBa_P0451 P0451]2009 Kit Plate 2 1N pSB1AK3

'''Parts needed to be built from scratch'''

'''prpBp''' [http://biocyc.org/ECOLI/NEW-IMAGE?type=OPERON&object=TU0-3602 Ecocyc]

(prpBp text version)

tgaaattaaa catttaattt tattaaggca attgtggcac accccttgct ttgtctttat caacgcaaat aacaagttga taaca

http://gcat.davidson.edu/GcatWiki/images/2/2f/PrpBp.JPG

'''prpB''' 2-methylisocitrate lyase in E.coli MG1655 [http://www.ncbi.nlm.nih.gov/sites/entrez?Db=gene&Cmd=ShowDetailView&TermToSearch=944990&ordinalpos=3&itool=EntrezSystem2.PEntrez.Gene.Gene_ResultsPanel.Gene_RVDocSum]http://gcat.davidson.edu/GcatWiki/images/6/6c/PrpB.JPG

(prpB text version)

atgTCTCTAC ACTCTCCAGG TAAAGCGTTT CGCGCTGCAC TGACTAAAGA AAATCCATTG

CAGATTGTTG GCACCATCAA CGCTAATCAT GCGCTGTTGG CGCAGCGTGC CGGATATCAG

GCAATTTATC TTTCTGGCGG TGGCGTGGCG GCAGGTTCGC TGGGGCTGCC CGATCTCGGT

ATTTCTACCC TTGATGATGT GCTGACCGAC ATTCGCCGTA TCACCGACGT TTGTTCGCTG

CCGCTGCTGG TGGATGCGGA TATCGGTTTT GGTTCTTCGG CCTTTAACGT GGCGCGCACC

GTGAAATCGA TGATTAAAGC CGGTGCGGCA GGATTGCATA TTGAAGATCA GGTTGGTGCG

AAACGCTGCG GTCATCGTCC GAATAAAGCG ATCGTCTCGA AAGAAGAGAT GGTGGATCGG

ATCCGCGCGG CGGTGGATGC GAAAACCGAT CCTGATTTTG TGATCATGGC GCGCACCGAT

GCTCTGGCGG TAGAGGGGCT GGATGCGGCG ATCGAGCGTG CGCAGGCCTA TGTTGAAGCG

GGTGCCGAGA TGTTGTTCCC GGAGGCGATT ACCGAACTCG CCATGTACCG CCAGTTTGCC

GATGCGGTGC AGGTGCCGAT CCTCGCCAAC ATCACCGAAT TTGGTGCCAC GCCGCTGTTT

ACCACCGACG AATTACGCAG CGCCCATGTC GCAATGGCGC TGTACCCACT TTCAGCGTTC

CGCGCCATGA ACCGCGCCGC TGAACATGTC TACAACGTCC TGCGCCAGGA AGGCACGCAG

AAAAGCGTCA TCGACACCAT GCAGACCCGC AACGAGCTGT ACGAAAGCAT CAACTACTAC

CAGTACGAAG AGAAGCTCGA CAACCTGTTT GCCCGTAGCC AGGTGAAATA A

'''pLuxT''' [http://www.jbc.org/cgi/reprint/281/46/34775]
http://gcat.davidson.edu/GcatWiki/images/e/e3/LuxT_upstream.JPG

(pLuxT text version)

agcgagcagt gagctcagtg gagtcatcag ttaaatctgt agtgttgtat tccacatttt

cgtttactgc tttcgccagc tctagatact tggcaaattc cgcttgtagc aattcttttg

aaccagcgta aatcgtcacg tctgcaacgt catcgccttc tttgatgata aagccaattt

caccagcgca accacacgct tcacatactt gggtttcttc gtgttcaatg ctcataacga

atcctcactt aaaaaaggga ttatactcaa ctcgtttttt tttcaataga aataaatgga

acaacttgga taaaaaaata tcggtatgct atgttgtttg ttgagagggt taagcaagag

ataactttag attttgttaa tggacgatgt gatccgagtc aataatttac cggatttcac

cttttactta cttagttatc acagtgagtg agttgcacct agcaagattt tataagagta

tctcgagata aattattttt acataatcta tcgctatcaa tcggttcaaa atgcagatgt

aaatattaaa ctaatgtcat ttaatggtat taatttagtt gcatattctg aaattttgtt

ttgcttgaag acatgcatct gcaaacctgg tttacagtat ttcgtaataa aaagtaggct

tgtttatttc gaaaaagggt gttttctggc aacctattga gaacgatttt cattctgacc

tcagactttg agttagatgg atttacaaag agaagagcta aaatatgcca aagcgtagta

aagaagatac cgaaatcact atccagaaaa ttatggatgc cgttgttgat cagttgctta

ggctcggtta cgacaaaatg tcgtacacga cgttgagtca acagacaggt gtttcacgta

(Xba1 site at bp81)

'''frame shift suppressor 1''' CU'''GUCCU'''AA

'''frame shift suppressor 2''' UU'''ACUAG'''AC

prpBp and prpB have no EcoR1, Xba1, Spe1, or Pst1 sites. Source www.neb.com

== References ==

Powerpoint [[Image:Suppressorlogic3.ppt]]

Anderson et al. 2007. “Environmental signal integration by a modular AND gate” [http://www.pubmedcentral.nih.gov/picrender.fcgi?artid=1964800&blobtype=pdf]

luxO luxT [http://www.jbc.org/cgi/content/full/281/46/34775]

PprpB propionate [http://www.pubmedcentral.nih.gov/articlerender.fcgi?tool=pubmed&pubmedid=16269719]

434 repressor [http://www.pnas.org/content/83/24/9353.full.pdf+html] [http://www.jbc.org/cgi/content/full/273/37/24165]