Template:SBB10 ConstructionFiles AndrewS sbb14
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2) sbb14: PhiC31 Integrase ---------------------------------------------------------- PCR AS009-F/AS010-R on pBca9523-Bca1623 (980 bp, gp = A) PCR AS011-F/AS012-R on pBca9523-Bca1659 (938 bp, gp = B) ---------------------------------------------------------- PCR AS009-F/AS010-R on A+B (1874 bp, EcoRI/BamHI) Sub in pBjk2741-Bca1144 (EcoRI/BamHI, 910+2170, L) Product is pBjk2741-sbb14 {rbs.phiC31>} ---------------------------------------------------------- AS009-F Forward PCR of Part 1 of PhiC31 with RBS ccataGAATTCatgAGATCTACATACCGAGATCATATATAAGGAGGTACATatgGACACGTACGCGGGTGCTTACG AS010-R SOEing of Part 1 of PhiC31 cggtaaccctcaatcttcgtGGTCGGCGTGCCGTCCGGCTTCTT AS011-F SOEing of Part 2 of PhiC31 AAGAAGCCGGACGGCACGCCGACCacgaagattgagggttaccg AS012-R Reverse PCR of Part 2 of PhiC31 atcagGGATCCCGCCGCTACGTCTTCCGT
JCA Notes
- BglII site missing from the first oligo
- Otherwise correct
- Need to put part into Clotho
sbb14: PhiC31 Integrase ---------------------------------------------------------- PCR AS009-F/AS010-R on Bca1623 6-9 (976bp, gp=A) PCR AS011-F/CA1674 on Bca1559 7-7 (514bp, gp=B) PCR CA1675/AS012-R on Bca1559 7-1 (444bp, gp=C) ---------------------------------------------------------- PCR AS009-F/AS012-R on A+B+C (1868 bp, EcoRI/BamHI) Sub in pBjk2741-Bca1144 (EcoRI/BamHI, 910+2170, L) Product is pBjk2741-sbb14 {rbs.phiC31>} ---------------------------------------------------------- AS009-F Forward PCR of Part 1 of PhiC31 with RBS ccataGAATTCatgAGATCTCAGATCATATATAAGGAGGTACATatgGACACGTACGCGGGTGCTTACG AS010-R SOEing of Part 1 of PhiC31 cggtaaccctcaatcttcgtGGTCGGCGTGCCGTCCGGCTTCTT AS011-F SOEing of Part 2 of PhiC31 AAGAAGCCGGACGGCACGCCGACCacgaagattgagggttaccg CA1674 GGGCGTCGGCGCGCTCCGCAACAAGGTTCGCCCGTTCGCCGCTCTTCTCAG PCA assembly of phiCthreeprime (Bca1659) CA1675 TGCGGAGCGCGCCGACGCCCTGAACGCCCTTGAAGAGCTGTACGAAGACCG PCA assembly of phiCthreeprime (Bca1659) AS012-R Reverse PCR of Part 2 of PhiC31 atcagGGATCCCGCCGCTACGTCTTCCGT Part: GATCTCAGATCATATATAAGGAGGTACATatgGACACGTACGCGGGTGCTTACGACCGTCAGTCGCGCGAGCGCGAAAATTCGAGCGCAGCAAGCCCAGCGACACAGCGTAGCGCCAACGAAGACAAGGCGGCCGACCTCCAGCGCGAAGTCGAGCGCGACGGGGGCCGGTTCAGGTTCGTCGGGCATTTCAGCGAAGCGCCGGGCACGTCGGCGTTCGGGACGGCGGAGCGCCCGGAGTTCGAACGCATCCTGAACGAATGCCGCGCCGGGCGGCTCAACATGATCATTGTCTATGACGTGTCGCGCTTCTCGCGCCTCAAGGTCATGGACGCGATTCCGATTGTCTCGGAATTGCTCGCCCTGGGCGTGACGATTGTTTCCACTCAGGAAGGCGTCTTCCGGCAGGGAAACGTCATGGACCTGATTCACCTGATTATGCGGCTCGACGCGTCGCACAAAGAATCTTCGCTCAAGTCGGCGAAGATTCTCGACACGAAGAACCTCCAGCGCGAATTGGGCGGGTACGTCGGCGGGAAGGCGCCTTACGGCTTCGAGCTTGTTTCGGAGACGAAGGAGATCACGCGCAACGGCCGAATGGTCAATGTCGTCATCAACAAGCTTGCGCACTCGACCACTCCCCTTACCGGACCCTTCGAGTTCGAGCCCGACGTAATCCGGTGGTGGTGGCGTGAGATCAAGACGCACAAACACCTTCCCTTCAAGCCGGGCAGTCAAGCCGCCATTCACCCGGGCAGCATCACGGGGCTTTGTAAGCGCATGGACGCTGACGCCGTGCCGACCCGGGGCGAGACGATTGGGAAGAAGACCGCTTCAAGCGCCTGGGACCCGGCAACCGTTATGCGAATCCTTCGGGACCCGCGTATTGCGGGCTTCGCCGCTGAGGTGATCTACAAGAAGAAGCCGGACGGCACGCCGACCacgaagattgagggttaccgCATTCAGCGCGACCCGATCACGCTCCGGCCGGTCGAGCTTGATTGCGGACCGATCATCGAGCCCGCTGAGTGGTATGAGCTCCAGGCGTGGTTGGACGGCAGGGGGCGCGGCAAGGGGCTTTCCCGGGGGCAAGCCATTCTGTCCGCCATGGACAAGCTGTACTGCGAGTGTGGCGCCGTCATGACTTCGAAGCGCGGGGAAGAATCGATCAAGGACTCTTACCGCTGCCGTCGCCGGAAGGTGGTCGACCCGTCCGCACCTGGGCAGCACGAAGGCACGTGCAACGTCAGCATGGCGGCACTCGACAAGTTCGTTGCGGAACGCATCTTCAACAAGATCAGGCACGCCGAAGGCGACGAAGAGACGTTGGCGCTTCTGTGGGAAGCCGCCCGACGCTTCGGCAAGCTCACTGAGGCGCCTGAGAAGAGCGGCGAACGGGCGAACCTTGTTGCGGAGCGCGCCGACGCCCTGAACGCCCTTGAAGAGCTGTACGAAGACCGCGCGGCAGGCGCGTACGACGGACCCGTTGGCAGGAAGCACTTCCGGAAGCAACAGGCAGCGCTGACGCTCCGGCAGCAAGGGGCGGAAGAGCGGCTTGCCGAACTTGAAGCCGCCGAAGCCCCGAAGCTTCCCCTTGACCAATGGTTCCCCGAAGACGCCGACGCTGACCCGACCGGCCCTAAGTCGTGGTGGGGGCGCGCGTCAGTAGACGACAAGCGCGTGTTCGTCGGGCTCTTCGTAGACAAGATCGTTGTCACGAAGTCGACTACGGGCAGGGGGCAGGGAACGCCCATCGAGAAGCGCGCTTCGATCACGTGGGCGAAGCCGCCGACCGACGACGACGAAGACGACGCCCAGGACGGCACGGAAGACGTAGCGGCGG