IGEM:PennState/Labbook/GalenLynch/2007-6-28

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June 28th, 2007:9:45-5:00

continued from the 26th Lambda bacteriophage regions of interest:

Lambda control region (OR)

  • Contains OR1, OR2, and OR3, shown in reverse order (i.e. the leftmost binding region below is OR3)
  • Bi directional promoter
    • in the lambda phage, CI is to the left(PRM), cro is to the right(PR)
  • recognized by cro, CI
    • cro binding affinities: OR3, OR2 ~ OR1 (OR1 is slightly less)
    • CI binding affinites: OR1, OR2 ~ OR3 (OR3 is slightly less)
  • CI acts as a positive regulator for PRM when bound to OR2, but and a negative regulator for PR when bound to OR3

<html><pre>ACGTTA<font color=cyan>AATC </font><font color= blue>TA</font><font color=green>TCACCGCA AGGGATA</font>AA<font color=cyan>T ATC</font><font color = blue>TA</font><font color = green>ACACC GTGCGTG</font><font color =blue>TTG</font><font color=cyan> ACT</font>ATTT<font color=green>TAC CTCTGGCGGT </font><font color=blue>GATA</font><font color=cyan>AT</font>GGTT GC</pre></html> Cro and CI binding regions are depicted in green, -10 and -35 sequences in cyan, and shared regions are in blue

Primers

 Readable primer:ggaattcgcg gccgcttcta gag ACGTTAAATC TATCACCGC (left side)
 Primer sequence:ggaattcgcggccgcttctagagACGTTAAATCTATCACCGC
  • Mp: 57,77
 Readable primer:ctgcagcggc cgctactagt a GCAACCATTA TCACCGC (right side)
 Primer sequence:ctgcagcggccgctactagtaGCAACCATTATCACCGC
  • Mp: 58,78

Lambda Repressor (CI)

  • Forms dimers
    • Dimers associate to form tetramers, octamers etc
  • Binds to OL and OR operators in bacteriophage genome
  • Cleaved by activated RecA protein (activates with ssDNA exposure)

<html><pre><font color=green>ATG</font>AGCACAA AAAAGAAACC ATTAACACAA GAGCAGCTTG AGGACGCACG TCGCCTTAAA GCAATTTATG AAAAAAAGAA AAATGAACTT GGCTTATCCC AGGAATCTGT CGCAGACAAG ATGGGGATGG GGCAGTCAGG CGTTGGTGCT TTATTTAATG GCATCAATGC ATTAAATGCT TATAACGCCG CATTGCTTGC AAAAATTCTC AAAGTTAGCG TTGAAGAATT TAGCCCTTCA ATCGCCAGAG AAATCTACGA GATGTATGAA GCGGTTAGTA TGCAGCCGTC ACTTAGAAGT GAGTATGAGT ACCCTGTTTT TTCTCATGTT CAGGCAGGGA TGTTCTCACC TGAGCTTAGA ACCTTTACCA AAGGTGATGC GGAGAGATGG GTAAGCACAA CCAAAAAAGC CAGTGATTCT GCATTCTGGC TTGAGGTTGA AGGTAATTCC ATGACCGCAC CAACAGGCTC CAAGCCAAGC TTTCCTGACG GAATGTTAAT TCTCGTTGAC CCTGAGCAGG CTGTTGAGCC AGGTGATTTC TGCATAGCCA GACTTGGGGG TGATGAGTTT ACCTTCAAGA AACTGATCAG GGATAGCGGT CAGGTGTTTT TACAACCACT AAACCCACAG TACCCAATGA TCCCATGCAA TGAGAGTTGT TCCGTTGTGG GGAAAGTTAT CGCTAGTCAG TGGCCTGAAG AGACGTTTGG C<font color=red>TGA</font></pre></html> Start codon is in green and stop codon is in red

Primers

 Readable Primer:ggaattcgcg gccgcttcta gag ATGAGCACAA AAAAGAAACC (left side)
 Primer sequence:ggaattcgcggccgcttctagagATGAGCACAAAAAAGAAACC
  • Mp: 56,77
 Readable primer:ctgcagcggc cgctactagt a TCAGCCAAAC GTCTCTTC (right side)
 Primer sequence:ctgcagcggccgctactagtaTCAGCCAAACGTCTCTTC
  • Mp: 58,78

Lambda Repressor (CI) with degredation tag (LVA)

Part BBa_C0051 on MIT's parts registry

  • same as CI repressor, but with degredation tag on the N terminus to promote degredation

Lambda AntiRepressor (Cro)

  • Binds to OR and OL operators
  • transcriptional activator, repressor
  • favors lytic pathway in lambda bacteriophage

<html><pre><font color=green>ATG</font>GAACAAC GCATAACCCT GAAAGATTAT GCAATGCGCT TTGGGCAAAC CAAGACAGCT AAAGATCTCG GCGTATATCA AAGCGCGATC AACAAGGCCA TTCATGCAGG CCGAAAGATT TTTTTAACTA TAAACGCTGA TGGAAGCGTT TATGCGGAAG AGGTAAAGCC CTTCCCGAGT AACAAAAAAA CAACAGCA<font color=red>TA A</font></pre></html> Start codon is in green and stop codon is in red

Primers

 Readable primer:ggaattcgcg gccgcttcta gag ATGGAACAAC GCATAACC (left side)
 Primer sequence:ggaattcgcggccgcttctagagATGGAACAACGCATAACC
  • Mp: 57,78
 Readable primer:ctgcagcggc cgctactagt a TTATGCTGTT GTTTTTTTGT TACT (right side)
 Primer sequence:ctgcagcggccgctactagtaTTATGCTGTTGTTTTTTTGTTACT
  • Mp: 58,75