Difference between revisions of "Oligos to Build"

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!width="50"|Reverse
 
!width="50"|Reverse
 
|-
 
|-
−
|luxS|| [[Image:luxS_forward.jpg]] || [[Image:luxS_reverse.jpg]]  
+
|luxS|| [[Image:luxS_forward.jpg]] || [[Image:lux_s_rev.jpg]]  
 
|-
 
|-
 
|lacIQ1 Promoter|| [[Image:placiq1_forward.jpg]] || [[Image:lacIq1reversecomplement.jpg]]  
 
|lacIQ1 Promoter|| [[Image:placiq1_forward.jpg]] || [[Image:lacIq1reversecomplement.jpg]]  
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|LacI|| [[Image:LacIforwardprimer.jpg]] || [[Image:LacIreversecomplement.jpg]]  
 
|LacI|| [[Image:LacIforwardprimer.jpg]] || [[Image:LacIreversecomplement.jpg]]  
 
|-
 
|-
−
|LacI_X86 (Primers for step 1 of PCR)|| [[Image:LacIforwardprimer.jpg]] || [[Image:X86_X86I12reversecomplementPCR1.jpg]]  
+
|LacI_X86 (Primers for step 1 of PCR)|| [[Image:LacIforwardprimer.jpg]] || [[Image:X86_X86I12reversecomplement.jpg]]  
 
|-
 
|-
−
|LacI_I12X86|| [[Image:LacI_I12forwardprimer.jpg]] || ###
+
|LacI_I12X86 (Primers for step 1 of PCR)|| [[Image:LacI_I12forwardprimer.jpg]] || [[Image:X86_X86I12reversecomplement.jpg]]
 
|-
 
|-
 
|###|| ### || ###  
 
|###|| ### || ###  
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!width="50"|Reverse
 
!width="50"|Reverse
 
|-
 
|-
−
|lsrR|| [[Image:lsr_forward.jpg]] || [[Image:lsr_reverse.jpg]]  
+
|lsrR|| [[Image:lsr_forward.jpg]] || [[Image:lsr_r_rev.jpg]]  
 
|-
 
|-
−
|lsrK|| [[Image:lsrK_forward.jpg]] || [[Image:lsrK_reverse.jpg]]
+
|lsrK|| [[Image:lsrK_forward.jpg]] || [[Image:lsrk_rev.jpg]]
 
|-
 
|-
−
|plsr||[[Image:plsr_forward.jpg]]|| [[Image:plsr_reverse.jpg]]  
+
|plsr||[[Image:plsr_forward.jpg]]|| [[Image:pLsr.jpg]]  
 
|-
 
|-
 
|###|| ### || ###  
 
|###|| ### || ###  
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|}
 
|}
  
 +
'''Davidson XOR Promoters''':
 +
 +
{| border="1" cellpadding="2"
 +
!width="50"|Part
 +
!width="50"|Oligos
 +
!width="50"|Oligo Design
 +
|-
 +
|lpLas'|| [[Image:pLasOligos.jpg]] || [[Image:pLas' oligos diagram.jpg]]
 +
|-
 +
|pLasXOR|| [[Image:pLasXOR Oligos.jpg]] || [[Image:pLasXOR.jpg]]
 +
|}
  
 
== '''Sequences of Parts to Build''' ==
 
== '''Sequences of Parts to Build''' ==
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AGCGGCATGCATTTACGTTGACACCACCTTTCGCGGTATGGCATGATAGCGCCCGG <br>
 
AGCGGCATGCATTTACGTTGACACCACCTTTCGCGGTATGGCATGATAGCGCCCGG <br>
 
-These promoter sequences are taken from Glascock and Weickert 1998
 
-These promoter sequences are taken from Glascock and Weickert 1998
 +
 +
'''TrpR Operator Sequence Only''' <br>
 +
ATATGCTATCGTACTCTTTAGCGAGTACAACCGGGGGA

Latest revision as of 20:32, 20 June 2008

Lac System:

Part Forward Reverse
luxS LuxS forward.jpg Lux s rev.jpg
lacIQ1 Promoter Placiq1 forward.jpg LacIq1reversecomplement.jpg
lacIQ Promoter LacIqforwardprimer.jpg LacIqreversecomplement.jpg
LacI_I12 LacI I12forwardprimer.jpg LacI I12reversecomplement.jpg
LacI LacIforwardprimer.jpg LacIreversecomplement.jpg
LacI_X86 (Primers for step 1 of PCR) LacIforwardprimer.jpg X86 X86I12reversecomplement.jpg
LacI_I12X86 (Primers for step 1 of PCR) LacI I12forwardprimer.jpg X86 X86I12reversecomplement.jpg
### ### ###

Lsr System:

Part Forward Reverse
lsrR Lsr forward.jpg Lsr r rev.jpg
lsrK LsrK forward.jpg Lsrk rev.jpg
plsr Plsr forward.jpg PLsr.jpg
### ### ###
### ### ###
### ### ###
### ### ###
### ### ###

Davidson XOR Promoters:

Part Oligos Oligo Design
lpLas' PLasOligos.jpg PLas' oligos diagram.jpg
pLasXOR PLasXOR Oligos.jpg PLasXOR.jpg

Sequences of Parts to Build

LuxS Wildtype from MG1655


ATGCCGTTGT TAGATAGCTT CACAGTCGAT CATACCCGGA TGGAAGCGCC TGCAGTTCGG GTGGCGAAAA CAATGAACAC CCCGCATGGC GACGCAATCA CCGTGTTCGA TCTGCGCTTC TGCGTGCCGA ACAAAGAAGT GATGCCAGAA AGAGGGATCC ATACCCTGGA GCACCTGTTT GCTGGTTTTA TGCGTAACCA TCTTAACGGT AATGGTGTAG AGATTATCGA TATCTCGCCA ATGGGCTGCC GCACCGGTTT TTATATGAGT CTGATTGGTA CGCCAGATGA GCAGCGTGTT GCTGATGCCT GGAAAGCGGC AATGGAAGAC GTGCTGAAAG TGCAGGATCA GAATCAGATC CCGGAACTGA ACGTCTACCA GTGTGGCACT TACCAGATGC ACTCGTTGCA GGAAGCGCAG GATATTGCGC GTAGCATTCT GGAACGTGAC GTACGCATCA ACAGCAACGA AGAACTGGCA CTGCCGAAAG AGAAGTTGCA GGAACTGCAC ATCTAG

Map Position: 2,812,240 <- 2,812,755 (nucleotides)

Sequence Length: 516 bp / 171 AAs

Mutated LuxS

Nucleotide position 54 changed from A to T in order to destroy a PstI site; this is a sense mutation

Also changed stop codon from TAG to TAA (recommended in Registry Help)

ATGCCGTTGT TAGATAGCTT CACAGTCGAT CATACCCGGA TGGAAGCGCC TGCTGTTCGG GTGGCGAAAA CAATGAACAC CCCGCATGGC GACGCAATCA CCGTGTTCGA TCTGCGCTTC TGCGTGCCGA ACAAAGAAGT GATGCCAGAA AGAGGGATCC ATACCCTGGA GCACCTGTTT GCTGGTTTTA TGCGTAACCA TCTTAACGGT AATGGTGTAG AGATTATCGA TATCTCGCCA ATGGGCTGCC GCACCGGTTT TTATATGAGT CTGATTGGTA CGCCAGATGA GCAGCGTGTT GCTGATGCCT GGAAAGCGGC AATGGAAGAC GTGCTGAAAG TGCAGGATCA GAATCAGATC CCGGAACTGA ACGTCTACCA GTGTGGCACT TACCAGATGC ACTCGTTGCA GGAAGCGCAG GATATTGCGC GTAGCATTCT GGAACGTGAC GTACGCATCA ACAGCAACGA AGAACTGGCA CTGCCGAAAG AGAAGTTGCA GGAACTGCAC ATCTAA

LsrR Gene

ATGACAATCA ACGATTCGGC AATTTCAGAA CAGGGAATGT GTGAAGAAGA ACAGGTCGCG CGGATCGCGT GGTTTTACTA TCACGACGGG CTGACCCAGA GCGAGATCAG CGATCGTCTC GGCCTGACAC GTTTGAAAGT GTCGCGATTG CTGGAGAAAG GGCATCAGTC CGGCATTATT CGCGTACAGA TTAATTCTCG CTTTGAAGGC TGTCTGGAAT ATGAAACTCA ATTACGTCGT CAGTTTTCGC TGCAACATGT CCGGGTGATC CCTGGGCTTG CGGATGCTGA TGTCGGTGGG CGACTGGGGA TAGGCGCGGC GCATATGTTG ATGAGTTTAC TTCAACCACA ACAGATGCTG GCGATTGGTT TTGGCGAGGC AACCATGAAT ACGCTGCAAC GCTTAAGTGG TTTTATTTCG TCACAGCAAA TTCGCCTGGT CACGCTCTCC GGTGGCGTCG GTTCTTATAT GACGGGAATC GGGCAGCTTA ACGCGGCGTG CAGTGTGAAT ATTATTCCGG CTCCGTTGCG GGCATCCTCC GCTGACATTG CCCGTACGCT AAAAAATGAA AATTGCGTCA AAGATGTTCT GTTAGCCGCG CAAGCAGCGG ATGTGGCGAT TGTCGGCATT GGTGCTGTGA GTCAACAGGA CGATGCGACA ATCATTCGCT CCGGTTATAT CAGCCAGGGC GAACAGTTAA TGATTGGCCG AAAAGGGGCG GTTGGCGACA TTTTAGGCTA CTTTTTTGAT GCAAAAGGTG ACGTTGTCAC GAATATCAAA ATACATAACG AACTGATTGG CTTACCTTTA AGCGCGCTGA AGACCATACC CGTCCGGGTT GGCGTGGCAG GGGGAGAAAA TAAAGCCGAA GCAATTGCCG CTGCAATGAA AGGCGGTTAT ATCAACGCAC TGGTTACCGA TCAGGACACA GCAGCGGCGA TTTTACGTAG TTAA

Map Position: 1,598,312 <- 1,599,265 (nucleotides)

Sequence Length: 954bp/317 AAs

LsrK Gene

atgGCTCGAC TCTTTACCCT TTCAGAATCA AAGTACTACC TGATGGCGCT GGATGCAGGC ACCGGAAGTA TTCGGGCTGT GATATTCGAC CTGGAAGGCA ATCAAATAGC AGTGGGACAG GCGGAGTGGC GGCATCTGGC AGTACCGGAC GTTCCTGGTT CTATGGAATT TGATCTCAAC AAAAACTGGC AACTGGCGTG TGAGTGTATG CGCCAGGCGC TGCACAACGC CGGCATAGCC CCGGAGTATA TCGCTGCCGT TTCGGCATGT TCGATGCGTG AAGGCATTGT TTTATATAAT AATGAAGGAG CCCCGATCTG GGCCTGCGCC AATGTGGATG CCAGAGCGGC ACGCGAAGTT AGCGAACTTA AAGAACTGCA CAACAATACC TTTGAAAACG AAGTTTATCG CGCGACCGGA CAAACACTGG CTTTAAGTGC CATCCCCAGA TTACTTTGGC TGGCGCACCA TCGTTCCGAT ATTTACCGTC AGGCATCAAC CATCACCATG ATCAGCGACT GGCTGGCCTA TATGCTCAGC GGCGAACTGG CGGTGGATCC CTCTAACGCT GGCACCACGG GACTTCTTGA TCTAACCACC CGTGACTGGA AACCTGCATT GCTGGATATG GCTGGCCTAC GTGCCGATAT TCTTTCTCCT GTCAAAGAAA CCGGCACATT GCTGGGCGTG GTAAGTTCAC AAGCGGCGGA ACTCTGCGGT CTGAAGGCGG GCACTCCGGT GGTCGTTGGA GGAGGCGACG TGCAGCTTGG TTGCCTTGGG TTAGGCGTTG TGCGTCCGGC ACAAACCGCG GTTCTTGGCG GCACATTCTG GCAGCAAGTT GTAAATTTAG CCGCGCCGGT GACAGACCCA GAAATGAACG TGCGCGTTAA TCCTCATGTT ATTCCTGGCA TGGTACAAGC TGAATCTATA AGCTTTTTTA CCGGACTCAC CATGCGCTGG TTCCGCGATG CTTTCTGTGC CGAAGAAAAA CTGATTGCCG AACGTTTAGG CATCGACACC TATACGCTGC TGGAAGAGAT GGCCAGTCGG GTGCCGCCTG GGTCGTGGGG CGTAATGCCG ATCTTCTCCG ACAGAATGCG CTTTAAAACC TGGTATCACG CTGCGCCTTC CTTTATTAAC TTGTCCATTG ACCCGGATAA ATGTAACAAA GCGACATTGT TCCGTGCGCT GGAAGAAAAT GCGGCGATTG TATCAGCGTG TAACTTGCAG CAAATTGCTG ATTTCTCGAA TATTCATCCT TCATCGTTAG TCTTTGCAGG CGGAGGTTCA AAAGGGAAAT TATGGAGTCA AATTCTCGCT GATGTCTCGG GATTACCCGT CAATATTCCG GTGGTCAAAG AAGCCACTGC ATTAGGATGT GCCATTGCAG CTGGCGTCGG TGCCGGAATT TTTTCATCAA TGGCAGAAAC CGGAGAACGC CTGGTTCGCT GGGAACGGAC GCACACACCA GACCCGGAAA AGCATGAACT TTATCAGGAT TCACGCGATA AGTGGCAGGC AGTTTATCAG GATCAGCTGG GGCTGGTTGA TCATGGACTG ACGACGTCGT TATGGAAAGC GCCTGGGTTA TAG

Map Position: 1,596,641 <- 1,598,233 (nucleotides)

Sequence Length: 1593 bp / 530 AAs

lsr promoter sequence
Plsr.jpg

LacI wild-type gene sequence (ORF begins at the first amino acid)
ATGAAACCAGTAACGTTATACGATGTCGCAGAGTATGCCGGTGTCTCTTATCAGACCGTTTCCCGCGTGGTGAACCAGGCCAGCCACGTTTCTGCGAAAACGCGGGAAAAAGTGGAAGC
GGCGATGGCGGAGCTGAATTACATTCCCAACCGCGTGGCACAACAACTGGCGGGCAAACAGTCGTTGCTGATTGGCGTTGCCACCTCCAGTCTGGCCCTGCACGCGCCGTCGCAA
ATTGTCGCGGCGATTAAATCTCGCGCCGATCAACTGGGTGCCAGCGTGGTGGTGTCGATGGTAGAACGAAGCGGCGTCGAAGCCTGTAAAGCGGCGGTGCACAATCTTCTCGCGC
AACGCGTCAGTGGGCTGATCATTAACTATCCGCTGGATGACCAGGATGCCATTGCTGTGGAAGCTGCCTGCACTAATGTTCCGGCGTTATTTCTTGATGTCTCTGACCAGACACCATCA
ACAGTATTATTTTCTCCCATGAAGACGGTACGCGACTGGGCGTGGAGCATCTGGTCGCATTGGGTCACCAGCAAATCGCGCTGTTAGCGGGCCCATTAAGTTCTGTCTCGGCG
CGTCTGCGTCTGGCTGGCTGGCATAAATATCTCACTCGCAATCAAATTCAGCCGATAGCGGAACGGGAAGGCGACTGGAGTGCCATGTCCGGTTTTCAACAAACCATGCAAATGC
TGAATGAGGGCATCGTTCCCACTGCGATGCTGGTTGCCAACGATCAGATGGCGCTGGGCGCAATGCGCGCCATTACCGAGTCCGGGCTGCGCGTTGGTGCGGATATCTCGGTAGT
GGGATACGACGATACCGAAGACAGCTCATGTTATATCCCGCCGTTAACCACCATCAAACAGGATTTTCGCCTGCTGGGGCAAACCAGCGTGGACCGCTTGCTGCAACTCTCTCAG
GGCCAGGCGGTGAAGGGCAATCAGCTGTTGCCCGTCTCACTGGTGAAAAGAAAAACCACCCTGGCGCCCAATACGCAAACCGCCTCTCCCCGCGCGTTGGCCGATTCATTAATGC
AGCTGGCACGACAGGTTTCCCGACTGGAAAGCGGGCAGTGA

LacI_I12 Mutation (amino acid 3 is changed from CCA to TAT)
ATGAAATATGTAACGTTATACGATGTCGCAGAGTATGCCGGTGTCTCTTATCAGACCGTTTCCCGCGTGGTGAACCAGGCCAGCCACGTTTCTGCGAAAACGCGGGAAAAAGTGGAAGC
GGCGATGGCGGAGCTGAATTACATTCCCAACCGCGTGGCACAACAACTGGCGGGCAAACAGTCGTTGCTGATTGGCGTTGCCACCTCCAGTCTGGCCCTGCACGCGCCGTCGCAA
ATTGTCGCGGCGATTAAATCTCGCGCCGATCAACTGGGTGCCAGCGTGGTGGTGTCGATGGTAGAACGAAGCGGCGTCGAAGCCTGTAAAGCGGCGGTGCACAATCTTCTCGCGC
AACGCGTCAGTGGGCTGATCATTAACTATCCGCTGGATGACCAGGATGCCATTGCTGTGGAAGCTGCCTGCACTAATGTTCCGGCGTTATTTCTTGATGTCTCTGACCAGACACCC
ATCAACAGTATTATTTTCTCCCATGAAGACGGTACGCGACTGGGCGTGGAGCATCTGGTCGCATTGGGTCACCAGCAAATCGCGCTGTTAGCGGGCCCATTAAGTTCTGTCTCGGC
GCGTCTGCGTCTGGCTGGCTGGCATAAATATCTCACTCGCAATCAAATTCAGCCGATAGCGGAACGGGAAGGCGACTGGAGTGCCATGTCCGGTTTTCAACAAACCATGCAAATG
CTGAATGAGGGCATCGTTCCCACTGCGATGCTGGTTGCCAACGATCAGATGGCGCTGGGCGCAATGCGCGCCATTACCGAGTCCGGGCTGCGCGTTGGTGCGGATATCTCGGTAG
TGGGATACGACGATACCGAAGACAGCTCATGTTATATCCCGCCGTTAACCACCATCAAACAGGATTTTCGCCTGCTGGGGCAAACCAGCGTGGACCGCTTGCTGCAACTCTCTCA
GGGCCAGGCGGTGAAGGGCAATCAGCTGTTGCCCGTCTCACTGGTGAAAAGAAAAACCACCCTGGCGCCCAATACGCAAACCGCCTCTCCCCGCGCGTTGGCCGATTCATTAATG
CAGCTGGCACGACAGGTTTCCCGACTGGAAAGCGGGCAGTGA

LacI_X86 Mutation (amino acid 61 is changed from TCG to CTG)
ATGAAACCAGTAACGTTATACGATGTCGCAGAGTATGCCGGTGTCTCTTATCAGACCGTTTCCCGCGTGGTGAACCAGGCCAGCCACGTTTCTGCGAAAACGCGGGAAAAAGTGGAAGC
GGCGATGGCGGAGCTGAATTACATTCCCAACCGCGTGGCACAACAACTGGCGGGCAAACAGCTGTTGCTGATTGGCGTTGCCACCTCCAGTCTGGCCCTGCACGCGCCGTCGCAA
ATTGTCGCGGCGATTAAATCTCGCGCCGATCAACTGGGTGCCAGCGTGGTGGTGTCGATGGTAGAACGAAGCGGCGTCGAAGCCTGTAAAGCGGCGGTGCACAATCTTCTCGCGC
AACGCGTCAGTGGGCTGATCATTAACTATCCGCTGGATGACCAGGATGCCATTGCTGTGGAAGCTGCCTGCACTAATGTTCCGGCGTTATTTCTTGATGTCTCTGACCAGACACCC
ATCAACAGTATTATTTTCTCCCATGAAGACGGTACGCGACTGGGCGTGGAGCATCTGGTCGCATTGGGTCACCAGCAAATCGCGCTGTTAGCGGGCCCATTAAGTTCTGTCTCGGC
GCGTCTGCGTCTGGCTGGCTGGCATAAATATCTCACTCGCAATCAAATTCAGCCGATAGCGGAACGGGAAGGCGACTGGAGTGCCATGTCCGGTTTTCAACAAACCATGCAAATG
CTGAATGAGGGCATCGTTCCCACTGCGATGCTGGTTGCCAACGATCAGATGGCGCTGGGCGCAATGCGCGCCATTACCGAGTCCGGGCTGCGCGTTGGTGCGGATATCTCGGTAG
TGGGATACGACGATACCGAAGACAGCTCATGTTATATCCCGCCGTTAACCACCATCAAACAGGATTTTCGCCTGCTGGGGCAAACCAGCGTGGACCGCTTGCTGCAACTCTCTCA
GGGCCAGGCGGTGAAGGGCAATCAGCTGTTGCCCGTCTCACTGGTGAAAAGAAAAACCACCCTGGCGCCCAATACGCAAACCGCCTCTCCCCGCGCGTTGGCCGATTCATTAATG
CAGCTGGCACGACAGGTTTCCCGACTGGAAAGCGGGCAGTGA

LacI_I12_X86 Mutation
ATGAAATATGTAACGTTATACGATGTCGCAGAGTATGCCGGTGTCTCTTATCAGACCGTTTCCCGCGTGGTGAACCAGGCCAGCCACGTTTCTGCGAAAACGCGGGAAAAAGTGGAAGCGGCGATGGCGGAG
CTGAATTACATTCCCAACCGCGTGGCACAACAACTGGCGGGCAAACAGCTGTTGCTGATTGGCGTTGCCACCTCCAGTCTGGCCCTGCACGCGCCGTCGCAAATTGTCGCGGCGATTAAATCTCGCGCC
GATCAACTGGGTGCCAGCGTGGTGGTGTCGATGGTAGAACGAAGCGGCGTCGAAGCCTGTAAAGCGGCGGTGCACAATCTTCTCGCGCAACGCGTCAGTGGGCTGATCATTAACTATCCGCTGGATGAC
CAGGATGCCATTGCTGTGGAAGCTGCCTGCACTAATGTTCCGGCGTTATTTCTTGATGTCTCTGACCAGACACCCATCAACAGTATTATTTTCTCCCATGAAGACGGTACGCGACTGGGCGTGGAGCATC
TGGTCGCATTGGGTCACCAGCAAATCGCGCTGTTAGCGGGCCCATTAAGTTCTGTCTCGGCGCGTCTGCGTCTGGCTGGCTGGCATAAATATCTCACTCGCAATCAAATTCAGCCGATAGCGGAACGGG
AAGGCGACTGGAGTGCCATGTCCGGTTTTCAACAAACCATGCAAATGCTGAATGAGGGCATCGTTCCCACTGCGATGCTGGTTGCCAACGATCAGATGGCGCTGGGCGCAATGCGCGCCATTACCGAGT
CCGGGCTGCGCGTTGGTGCGGATATCTCGGTAGTGGGATACGACGATACCGAAGACAGCTCATGTTATATCCCGCCGTTAACCACCATCAAACAGGATTTTCGCCTGCTGGGGCAAACCAGCGTGGACC
GCTTGCTGCAACTCTCTCAGGGCCAGGCGGTGAAGGGCAATCAGCTGTTGCCCGTCTCACTGGTGAAAAGAAAAACCACCCTGGCGCCCAATACGCAAACCGCCTCTCCCCGCGCGTTGGCCGATTCAT
TAATGCAGCTGGCACGACAGGTTTCCCGACTGGAAAGCGGGCAGTGA

LacI Promoter
CGTTGACACCATCGAATGGCGCAAAACCTTTCGCGGTATGGCATGATAGCGCCCGG

LacIQ Promoter
CGTTGACACCATCGAATGGTGCAAAACCTTTCGCGGTATGGCATGATAGCGCCCGG

LacIQ1 Promoter
AGCGGCATGCATTTACGTTGACACCACCTTTCGCGGTATGGCATGATAGCGCCCGG
-These promoter sequences are taken from Glascock and Weickert 1998

TrpR Operator Sequence Only
ATATGCTATCGTACTCTTTAGCGAGTACAACCGGGGGA

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