IGEM:IMPERIAL/2009/Primers

=Primers to be used for PCR=

Considerations for primer design http://www.premierbiosoft.com/tech_notes/PCR_Primer_Design.html

Use this website to calculate temperatures and modify primers and this one for reverse complement.

RcsB Sequence:

ATGAACAATATGAACGTAATTATTGCCGATGACCATCCGATAGTCTTGTTCGGTATTCGCAAATCACTTG AGCAAATTGAGTGGGTGAATGTTGTCGGCGAATTTGAAGACTCTACAGCACTGATCAACAACCTGCCGAA ACTGGATGCGCATGTGTTGATTACCGATCTCTCCATGCCTGGCGATAAGTACGGCGATGGCATTACCTTA ATCAAGTACATCAAGCGCCATTTCCCAAGCCTGTCGATCATTGTTCTGACTATGAACAACAACCCGGCGA TTCTTAGTGCGGTATTGGATCTGGATATCGAAGGGATCGTGCTGAAACAAGGTGCACCGACCGATCTGCC GAAAGCTCTCGCCGCGCTGCAGAAAGGGAAGAAATTTACCCCGGAAAGCGTTTCTCGCCTGTTGGAAAAA ATCAGTGCTGGTGGTTACGGTGACAAGCGTCTCTCGCCAAAAGAGAGTGAAGTTCTGCGCCTGTTTGCGG AAGGCTTCCTGGTGACCGAGATCGCTAAAAAGCTGAACCGCAGTATTAAAACCATCAGTAGCCAGAAGAA ATCTGCGATGATGAAGCTGGGTGTCGAGAACGATATCGCCCTGCTGAATTATCTCTCTTCAGTGACCTTA AGTCCGGCAGATAAAGACTAA

Forward: GCTCTAGATGAACAATATGAACGTAATTATTG      53.1C(without) and 61.8(with) Backward: GGACTAGTATTAGTCTTTATCTGCCGG      51.4C(without) and 63.4(with) Sequence Length: 651bp Strains to use:DH10B/TOP10 OtsB Sequence:

ATGACAGAACCGTTAACCGAAACCCCTGAACTATCCGCGAAATATGCCTGGTTTTTTGATCTTGATG GAACGCTGGCGGAAATCAAACCGCATCCCGATCAGGTCGTCGTGCCTGACAATATTCTGCAAGGACT ACAGCTACTGGCAACCGCAAGTGATGGTGCATTGGCATTGATATCAGGGCGCTCAATGGTGGAGCTT GACGCACTGGCAAAACCTTATCGCTTCCCGTTAGCGGGCGTGCATGGGGCGGAGCGCCGTGACATCA ATGGTAAAACACATATCGTTCATCTGCCGGATGCGATTGCGCGTGATATTAGCGTGCAACTGCATAC AGTCATCGCTCAGTATCCCGGCGCGGAGCTGGAGGCGAAAGGGATGGCTTTTGCGCTGCATTATCGT CAGGCTCCGCAGCATGAAGACGCATTAATGACATTAGCGCAACGTATTACTCAGATCTGGCCACAAA TGGCGTTACAGCAGGGAAAGTGTGTTGTCGAGATCAAACCGAGAGGTACCAGTAAAGGTGAGGCAAT TGCAGCTTTTATGCAGGAAGCTCCCTTTATCGGGCGAACGCCCGTATTTCTGGGCGATGATTTAACC GATGAATCTGGCTTCGCAGTCGTTAACCGACTGGGCGGAATGTCAGTAAAAATTGGCACAGGTGCAA CTCAGGCATCATGGCGACTGGCGGGTGTGCCGGATGTCTGGAGCTGGCTTGAAATGATAACCACCGC ATTACAACAAAAAAGAGAAAATAACAGGAGTGATGACTATGAGTCGTTTAGTCGTAGTATCTAA Forward:	GCTCTAGATGACAGAACCGTTAACCG      54.5C (without) & 64.8C (with) Backward:	GGACTAGTATTAGATACTACGACTAAACGAC      54.7C (without) & 64.2C (with) Sequence Length: 805bp Strains to use:BL21(DE3) B3023 Sequence: ATGACAAACCTGACACTGGATGTAAACATTATCGATTTCCCATCAATACCTGTGGCGATGTTGCCG CACCGCTGTAGCCCTGAATTGCTCAACTACAGCGTGGCGAAATTTATCATGTGGCGTAAAGAGACG GGGCTTTCTCCTGTTAACCAAAGCCAGACTTTTGGCGTCGCCTGGGACGACCCTGCCACCACCGCA CCGGAAGCGTTTCGCTTTGATATCTGCGGCAGCGTTAGCGAACCGATTCCCGATAATCGTTATGGT GTGAGCAATGGTGAACTTACCGGTGGACGTTATGCCGTGGCCCGCCACGTTGGCGAGCTGGACGAT ATTTCACACACGGTATGGGGCATCATTCGCCACTGGCTGCCTGCAAGCGGCGAGAAAATGCGTAAA GCACCGATTCTGTTTCACTACACCAATCTTGCCGAAGGGGTGACAGAGCAGCGACTGGAAACGGAT GTTTATGTGCCGTTGGCGTGA

Forward: GCTCTAGATGACAAACCTGACACTG      51.4C(without) and 63.0(with) Backward: GGACTAGTATCACGCCAACGGCAC      53.3C(without) and 66.1C(with) Sequence Length: 483bp Strains to use:BL21(DE3) WaaL Sequence:

ATGCTAACATCCTTTAAACTTCATTCATTGAAACCTTACACTCTGAAATCATCAATGATTTTAGAG ATAATAACTTATATATTATGTTTTTTTTCAATGATAATTGCATTCGTCGATAATACTTTCAGCATA AAAATATATAATATCACTGCTATAGTTTGCTTATTGTCACTAATTTTACGTGGCAGACAAGAAAATT ATAATATAAAAAACCTTATTCTTCCCCTTTCTATATTTTTAATAGGCTTGCTTGATTTAATTTGGTA TTCTGCGTTTAAAGTAGATAATTCGCCATTTCGTGCTACTTACCATAGTTATTTAAATACTGCCAAA ATATTTATATTTGGTTCTTTTATTGTTTTCTTGACACTAACTAGCCAGCTAAAATCAAAAAAAGAGA GTGTATTATACACTTTGTATTCTCTGTCATTTCTAATTGCTGGATATGCAATGTATATTAATAGCAT TCATGAAAATGACCGCATTTCTTTTGGTGTAGGAACGGCAACAGGAGCAGCATATTCAACAATGCTA ATAGGGATAGTTAGTGGCGTTGCGATTCTTTATACTAAGAAAAATCATCCTTTTTTATTTTTATTAA ATAGTTGCGCGGTACTTTATGTTCTGGCGCTAACACAAACCAGAGCAACCCTACTCCTGTTCCCTAT AATTTGTGTTGCTGCATTAATAGCTTATTATAATAAATCACCCAAGAAATTCACTTCCTCTATTGTT CTACTAATTGCTATATTAGCTAGCATTGTTATTATATTTAATAAACCAATACAGAATCGCTATAATG AAGCATTAAATGACTTAAACAGTTATACCAATGCTAATAGTGTTACTTCCCTAGGTGCAAGACTGGC AATGTACGAAATTGGTTTAAATATATTCATAAAGTCACCTTTTTCATTTAGATCAGCAGAGTCACGC GCTGAAAGTATGAATTTGTTAGTTGCAGAACACAATAGGCTAAGAGGGGCATTGGAGTTTTCTAACG TACATCTACATAATGAGATAATTGAAGCAGGGTCACTGAAAGGTCTGATGGGAATTTTTTCCACACT TTTCCTCTATTTTTCACTATTTTATATAGCATATAAAAAACGAGCTTTGGGTTTGTTGATATTAACG CTTGGCATTGTGGGGATTGGACTCAGTGATGTGATCATATGGGCACGCAGCATTCCAATTATCATTA TATCCGCTATAGTCCTCTTACTCGTCATTAATAATCGTAACAATACAATTAATTAA

Forward: GCTCTAGATGCTAACATCCTTTAAACTTC      52.8C(without) and 62.4(with) Backward: GGACTAGTATTAATTAATTGTATTGTTACGATTATTAATG      54.9C(without) and 62.3(with) Sequence Length: 1260bp Strains to use:DH10B/TOP10 OtsA Sequence:

ATGAGTCGTTTAGTCGTAGTATCTAACCGGATTGCACCACCAGACGAGCACGCCGCCAGTGCCGGTGGCCTTGC CGTTGGCATACTGGGGGCACTGAAAGCCGCAGGCGGACTGTGGTTTGGCTGGAGTGGTGAAACAGGGAATGAGG ATCAGCCGCTAAAAAAGGTGAAAAAAGGTAACATTACGTGGGCCTCTTTTAACCTCAGCGAACAGGACCTTGAC GAATACTACAACCAATTCTCCAATGCCGTTCTCTGGCCCGCTTTTCATTATCGGCTCGATCTGGTGCAATTTCA GCGTCCTGCCTGGGACGGCTATCTACGCGTAAATGCGTTGCTGGCAGATAAATTACTGCCGCTGTTGCAAGACG ATGACATTATCTGGATCCACGATTATCACCTGTTGCCATTTGCGCATGAATTACGCAAACGGGGAGTGAATAAT CGCATTGGTTTCTTTCTGCATATTCCTTTCCCGACACCGGAAATCTTCAACGCGCTGCCGACATATGACACCTT GCTTGAACAGCTTTGTGATTATGATTTGCTGGGTTTCCAGACAGAAAACGATCGTCTGGCGTTCCTGGATTGTC TTTCTAACCTGACCCGCGTCACGACACGTAGCGCAAAAAGCCATACAGCCTGGGGCAAAGCATTTCGAACAGAA GTCTACCCGATCGGCATTGAACCGAAAGAAATAGCCAAACAGGCTGCCGGGCCACTGCCGCCAAAACTGGCGCA ACTTAAAGCGGAACTGAAAAACGTACAAAATATCTTTTCTGTCGAACGGCTGGATTATTCCAAAGGTTTGCCAG AGCGTTTTCTCGCCTATGAAGCGTTGCTGGAAAAATATCCGCAGCATCATGGTAAAATTCGTTATACCCAGATT GCACCAACGTCGCGTGGTGATGTGCAAGCCTATCAGGATATTCGTCATCAGCTCGAAAATGAAGCTGGACGAAT TAATGGTAAATACGGGCAATTAGGCTGGACGCCGCTTTATTATTTGAATCAGCATTTTGACCGTAAATTACTGA TGAAAATATTCCGCTACTCTGACGTGGGCTTAGTGACGCCACTGCGTGACGGGATGAACCTGGTAGCAAAAGAG TATGTTGCTGCTCAGGACCCAGCCAATCCGGGCGTTCTTGTTCTTTCGCAATTTGCGGGAGCGGCAAACGAGTT AACGTCGGCGTTAATTGTTAACCCCTACGATCGTGACGAAGTTGCAGCTGCGCTGGATCGTGCATTGACTATGT CGCTGGCGGAACGTATTTCCCGTCATGCAGAAATGCTGGACGTTATCGTGAAAAACGATATTAACCACTGGCAG GAGTGCTTCATTAGCGACCTAAAGCAGATAGTTCCGCGAAGCGCGGAAAGCCAGCAGCGCGATAAAGTTGCTAC CTTTCCAAAGCTTGCGTAG

Forward: GCTCTAGATGAGTCGTTTAGTCGTAG      52.4C(without) and 63.2(with) Backward: GGACTAGTACTACGCAAGCTTTGGAAAG      54.5C(without) and 65.1(with) Sequence Length: 1425bp Strain to use:BL21(DE3) PAH Sequence:

ATGTCCACTGCGGTCCTGGAAAACCCAGGCTTGGGCAGGAAACTCTCTGACTTTGGACAGGAAACAAGCTATAT TGAAGACAACTGCAATCAAAATGGTGCCATATCACTGATCTTCTCACTCAAAGAAGAAGTTGGTGCATTGGCCA AAGTATTGCGCTTATTTGAGGAGAATGATGTAAACCTGACCCACATTGAATCTAGACCTTCTCGTTTAAAGAAA GATGAGTATGAATTTTTCACCCATTTGGATAAACGTAGCCTGCCTGCTCTGACAAACATCATCAAGATCTTGAG GCATGACATTGGTGCCACTGTCCATGAGCTTTCACGAGATAAGAAGAAAGACACAGTGCCCTGGTTCCCAAGAA CCATTCAAGAGCTGGACAGATTTGCCAATCAGATTCTCAGCTATGGAGCGGAACTGGATGCTGACCACCCTGGT TTTAAAGATCCTGTGTACCGTGCAAGACGGAAGCAGTTTGCTGACATTGCCTACAACTACCGCCATGGGCAGCC CATCCCTCGAGTGGAATACATGGAGGAAGAAAAGAAAACATGGGGCACAGTGTTCAAGACTCTGAAGTCCTTGT ATAAAACCCATGCTTGCTATGAGTACAATCACATTTTTCCACTTCTTGAAAAGTACTGTGGCTTCCATGAAGAT AACATTCCCCAGCTGGAAGACGTTTCTCAATTCCTGCAGACTTGCACTGGTTTCCGCCTCCGACCTGTGGCTGG CCTGCTTTCCTCTCGGGATTTCTTGGGTGGCCTGGCCTTCCGAGTCTTCCACTGCACACAGTACATCAGACATG GATCCAAGCCCATGTATACCCCCGAACCTGACATCTGCCATGAGCTGTTGGGACATGTGCCCTTGTTTTCAGAT CGCAGCTTTGCCCAGTTTTCCCAGGAAATTGGCCTTGCCTCTCTGGGTGCACCTGATGAATACATTGAAAAGCT CGCCACAATTTACTGGTTTACTGTGGAGTTTGGGCTCTGCAAACAAGGAGACTCCATAAAGGCATATGGTGCTG GGCTCCTGTCATCCTTTGGTGAATTACAGTACTGCTTATCAGAGAAGCCAAAGCTTCTCCCCCTGGAGCTGGAG AAGACAGCCATCCAAAATTACACTGTCACGGAGTTCCAGCCCCTGTATTACGTGGCAGAGAGTTTTAATGATGC CAAGGAGAAAGTAAGGAACTTTGCTGCCACAATACCTCGGCCCTTCTCAGTTCGCTACGACCCATACACCCAAA GGATTGAGGTCTTGGACAATACCCAGCAGCTTAAGATTTTGGCTGATTCCATTAACAGTGAAATTGGAATCCTT TGCAGTGCCCTCCAGAAAATAAAGTAA

Forward:	GCTCTAGATGTCCACTGCGGTCC      54.3C (without) & 66.0C (with) Backward:	GGACTAGTATTACTTTATTTTCTGGAGGGC      54.0C (without) & 64.0C (with) Sequence Length: 1359bp Cellulase Sequence:

ATGAAAAAAATTGTTTCTTTGGTTTGTGTGCTTGTGATGCTGGTAAGCATCTTAGGCTCGTTTTCAGTCGTA GCGGCATCACCGGTAAAAGGCTTTCAGGTATCGGGAACAAAGCTTTTGGATGCAAGCGGAAACGAGCTTGTA ATGAGGGGCATGCGTGATATTTCAGCAATAGATTTGGTTAAAGAAATAAAAATCGGATGGAATTTGGGAAAT ACTTTGGATGCTCCTACAGAGACTGCCTGGGGAAATCCAAGGACAACCAAGGCAATGATAGAAAAGGTAAGG GAAATGGGCTTTAATGCCGTCAGAGTGCCTGTTACCTGGGATACGCACATCGGACCTGCTCCGGACTATAAA ATTGACGAAGCATGGCTGAACAGAGTTGAGGAAGTGGTAAACTATGTTCTTGACTGCGGTATGTACGCGATC ATAAATGTTCACCATGACAATACATGGATTATACCTACATATGCCAATGAGCAAAGGAGTAAAGAAAAACTT GTAAAAGTTTGGGAACAAATAGCAACCCGTTTTAAAGATTATGACGACCATTTGTTGTTTGAGACAATGAAC GAACCGAGAGAAGTAGGTTCACCTATGGAATGGATGGGCGGAACGTATGAAAACCGAGATGTGATAAACAGA TTTAATTTGGCGGTTGTTAATACCATCAGAGCAAGCGGCGGAAATAACGATAAAAGATTCATACTGGTTCCG ACCAATGCGGCAACCGGCCTGGATGTTGCATTAAACGACCTTGTCATTCCGAACAATGACAGCAGAGTCATA GTATCCATACATGCTTATTCACCGTATTTCTTTGCTATGGATGTCAACGGAACTTCATATTGGGGAAGTGAC TATGACAAGGCTTCTCTTACAAGTGAACTTGATGCTATTTACAACAGATTTGTGAAAAACGGAAGGGCTGTA ATTATCGGAGAATTCGGAACCATTGACAAGAACAACCTGTCTTCAAGGGTGGCTCATGCCGAGCACTATGCA AGAGAAGCAGTTTCAAGAGGAATTGCTGTTTTCTGGTGGGATAACGGCTATTACAATCCGGGTGATGCAGAG ACTTATGCATTGCTGAACAGAAAAACTCTCTCATGGTATTATCCTGAAATTGTCCAGGCTCTTATGAGAGGT GCCGGCGTTGAACCTTTAGTTTCACCGACTCCTACACCTACATTAATGCCGACCCCCTCGCCCACGGTGACA GCAAATATTTTGTACGGTGACGTAAACGGGGACGGAAAAATAAATTCTACAGACTGTACAATGCTAAAGAGA TATATTTTGCGTGGCATAGAAGAATTCCCAAGTCCTAGCGGAATTATAGCCGCTGACGTAAATGCGGATCTG AAAATCAATTCCACCGACTTGGTATTGATGAAAAAATATCTACTGCGCTCAATAGACAAATTTCCTGCGGAG GATTCTCAAACACCTGATGAAGACAATCCGGGCATTTTGTATAACGGAAGATTCGATTTTTCAGATCCGAAC GGTCCGAAATGCGCCTGGTCCGGCAGCAATGTTGAGCTGAATTTTTACGGCACGGAAGCAAGTGTGACTATC AAATCCGGCGGTGAGAACTGGTTCCAGGCTATTGTAGACGGCAATCCTCTTCCTCCTTTTTCGGTTAACGCT ACTACCTCTACCGTAAAGCTTGTAAGCGGTCTTGCAGAAGGAGCTCATCATCTTGTATTGTGGAAGAGGACA GAGGCATCCTTGGGAGAAGTTCAGTTCCTTGGGTTTGATTTTGGTTCAGGAAAGCTTCTTGCCGCACCGAAG CCTTTGGAAAGAAAGATTGAGTTTATCGGAGACTCCATCACATGTGCATACGGAAATGAAGGAACAAGCAAG GAGCAGTCTTTTACACCGAAAAATGAAAACAGCTATATGTCTTATGCGGCAATTACAGCCCGTAATTTGAAT GCAAGTGCAAATATGATTGCGTGGTCCGGAATCGGACTTACCATGAACTACGGCGGAGCCCCCGGACCTCTT ATAATGGACCGTTATCCTTATACCCTTCCTTACAGCGGAGTCAGATGGGATTTTAGCAAATATGTGCCTCAG GTTGTTGTAATCAATCTTGGTACCAATGATTTTTCTACATCATTTGCAGATAAAACAAAGTTTGTAACGGCA TATAAAAACCTTATAAGTGAAGTTCGCAGGAACTATCCGGATGCCCATATATTCTGCTGTGTCGGTCCGATG CTTTGGGGAACGGGCCTGGATTTGTGCCGCAGTTATGTTACGGAAGTTGTAAATGATTGTAACAGAAGCGGG GATTTAAAGGTGTATTTTGTTGAGTTTCCGCAGCAGGACGGAAGCACCGGATACGGAGAAGACTGGCATCCA AGTATTGCCACCCACCAGCTGATGGCTGAGCGGCTTACTGCGGAAATAAAAAACAAGCTTGGATGGTAA

Forward:	GCTCTAGATGAAAAAAATTGTTTCTTTGGTTTG      53.8C (without) & 62.0C (with) Backward:	GGACTAGTATTACCATCCAAGCTTGTTTTTTATTTC      57.4C (without) & 64.9C (with) Sequence Length: 2445bp Dam Sequence:

ATGAAGAAAAATCGCGCTTTTTTGAAGTGGGCAGGGGGCAAGTATCCCCTGCTTGATGATATTAAA CGGCATTTGCCCAAGGGCGAATGTCTGGTTGAGCCTTTTGTAGGTGCCGGGTCGGTGTTTCTCAAC ACCGACTTTTCTCGTTATATCCTTGCCGATATCAATAGCGACCTGATCAGTCTCTATAACATTGTG AAGATGCGTACTGATGAGTACGTACAGGCCGCACGCGAGCTGTTTGTTCCCGAAACAAATTGCGCC GAGGTTTACTATCAGTTCCGCGAAGAGTTCAACAAAAGCCAGGATCCGTTCCGTCGGGCGGTACTG TTTTTATATTTGAACCGCTACGGTTACAACGGCCTGTGTCGTTACAATCTGCGCGGTGAGTTTAAC GTGCCGTTCGGCCGCTACAAAAAACCCTATTTCCCGGAAGCAGAGTTGTATCACTTCGCTGAAAAA GCGCAGAATGCCTTTTTCTATTGTGAGTCTTACGCCGATAGCATGGCGCGCGCAGATGATGCATCC GTCGTCTATTGCGATCCGCCTTATGCACCGCTGTCTGCGACCGCCAACTTTACGGCGTATCACACA AACAGTTTTACGCTTGAACAACAAGCGCATCTGGCGGAGATCGCCGAAGGTCTGGTTGAGCGCCAT ATTCCAGTGCTGATCTCCAATCACGATACGATGTTAACGCGTGAGTGGTATCAGCGCGCAAAATTG CATGTCGTCAAAGTTCGACGCAGTATAAGCAGCAACGGCGGCACACGTAAAAAGGTGGACGAACTG CTGGCTTTGTACAAACCAGGAGTCGTTTCACCCGCGAAAAAATAA

Forward: GCTCTAGATGAAGAAAAATCGCGCTTTTTTG      55.9C (without) & 64.2C (with) Backward: GGACTAGTATTATTTTTTCGCGGGTGAAAC      54.0C (without) & 64.0C (with) Sequence Length: 837bp Strain to use:BL21(DE3)