Gene replacement using the pKO3 vector
NOTE: The file is constructed from various GenBank files and is not
primary sequence and has not been confirmed.
Comments:
Sequence file of the gene replacement vector pKO3.
The vector has a temperature-sensitive pSC101 replicon which
which is functional at 30 C.
Hamiliton et. al. ,1989 J. Bacteriology 171: 4617-4622
Link, A.J., Phillips, D. and Church, G.M. (1997)
Methods for generating precise deletions and insertions in the
genome of wild-type Escherichia coli:
Application to open reading frame characterization.
J. Bacteriology 179: 6228-6237.
Current base coordinates:
221-880 Cm-R; chloramphenicol acetyltransferase
1119-1575 f1 (+) filamentous origin
2129-3550 Bacillus subtilis sacB gene
4139-4149 oriC homologous region
4204-4215 IHF binding site
4407-5357 RepA (temperature-sensitive ?)
Pko3.Mail Length: 5681 May 31, 1995 11:25 Type: N Check: 5438 ..
1 CCCTTTCGTC TTCGAATAAA TACCTGTGAC GGAAGATCAC TTCGCAGAAT
51 AAATAAATCC TGGTGTCCCT GTTGATACCG GGAAGCCCTG GGCCAACTTT
101 TGGCGAAAAT GAGACGTTGA TCGGCACGTA AGAGGTTCCA ACTTTCACCA
151 TAATGAAATA AGATCACTAC CGGGCGTATT TTTTGAGTTA TCGAGATTTT
201 CAGGAGCTAA GGAAGCTAAA ATGGAGAAAA AAATCACTGG ATATACCACC
251 GTTGATATAT CCCAATGGCA TCGTAAAGAA CATTTTGAGG CATTTCAGTC
301 AGTTGCTCAA TGTACCTATA ACCAGACCGT TCAGCTGGAT ATTACGGCCT
351 TTTTAAAGAC CGTAAAGAAA AATAAGCACA AGTTTTATCC GGCCTTTATT
401 CACATTCTTG CCCGCCTGAT GAATGCTCAT CCGGAATTCC GTATGGCAAT
451 GAAAGACGGT GAGCTGGTGA TATGGGATAG TGTTCACCCT TGTTACACCG
501 TTTTCCATGA GCAAACTGAA ACGTTTTCAT CGCTCTGGAG TGAATACCAC
551 GACGATTTCC GGCAGTTTCT ACACATATAT TCGCAAGATG TGGCGTGTTA
601 CGGTGAAAAC CTGGCCTATT TCCCTAAAGG GTTTATTGAG AATATGTTTT
651 TCGTCTCAGC CAATCCCTGG GTGAGTTTCA CCAGTTTTGA TTTAAACGTG
701 GCCAATATGG ACAACTTCTT CGCCCCCGTT TTCACCATGG GCAAATATTA
751 TACGCAAGGC GACAAGGTGC TGATGCCGCT GGCGATTCAG GTTCATCATG
801 CCGTTTGTGA TGGCTTCCAT GTCGGCAGAA TGCTTAATGA ATTACAACAG
851 TACTGCGATG AGTGGCAGGG CGGGGCGTAA TTTTTTTAAG GCAGTTATTG
901 GTGCCCTTAA ACGCCTGGTG CTACGCCTGA ATAAGTGATA ATAAGCGGAT
951 GAATGGCAGA AATTCGAAAG CAAATTCGAC CCGGTCGTCG GTTCAGGGCA
1001 GGGTCGTTAA ATAGCCGCTT ATGTCTATTG CTGGTTTANT CGGTACCCGG
1051 Ggatcgcggc cgcggaccgg atcctctaga gcggccgcGA TCCTCTAGAG
1101 TCGACCGGNG AATGGCGAAT GGGACGCGCC CTGTAGCGGC GCATTAAGCG
1151 CGGCGGGTGT GGTGGTTACG CGCAGCGTGA CCGCTACACT TGCCAGCGCC
1201 CTAGCGCCCG CTCCTTTCGC TTTCTTCCCT TCCTTTCTCG CCACGTTCGC
1251 CGGCTTTCCC CGTCAAGCTC TAAATCGGGG GCTCCCTTTA GGGTTCCGAT
1301 TTAGTGCTTT ACGGCACCTC GACCCCAAAA AACTTGATTA GGGTGATGGT
1351 TCACGTAGTG GGCCATCGCC CTGATAGACG GTTTTTCGCC CTTTGACGTT
1401 GGAGTCCACG TTCTTTAATA GTGGACTCTT GTTCCAAACT GGAACAACAC
1451 TCAACCCTAT CTCGGTCTAT TCTTTTGATT TATAAGGGAT TTTGCCGATT
1501 TCGGCCTATT GGTTAAAAAA TGAGCTGATT TAACAAAAAT TTAACGCGAA
1551 TTTTAACAAA ATATTAACGC TTACAATTTA GGTGGCACTT TTCGGGGAAA
1601 TGTGCGCGGA ACCCCTATTT GTTTATTTTT CTAAATACAT TCAAATATGT
1651 ATCCGCTCAT NNCANGATCC TTTTTAACCC ATCACATATA CCTGCCGTTC
1701 ACTATTATTT AGTGAAATGA GATATTATGA TATTTTCTGA ATTGTGATTA
1751 AAAAGGCAAC TTTATGCCCA TGCAACAGAA ACTATAAAAA ATACAGAGAA
1801 TGAAAAGAAA CAGATAGATT TTTTAGTTCT TTAGGCCCGT AGTCTGCAAA
1851 TCCTTTTATG ATTTTCTATC AAACAAAAGA GGAAAATAGA CCAGTTGCAA
1901 TCCAAACGAG AGTCTAATAG AATGAGGTCG AAAAGTAAAT CGCGCGGGTT
1951 TGTTACTGAT AAAGCAGGCA AGACCTAAAA TGTGTAAAGG GCAAAGTGTA
2001 TACTTTGGCG TCACCCCTTA CATATTTTAG GTCTTTTTTT ATTGTGCGTA
2051 ACTAACTTGC CATCTTCAAA CAGGAGGGCT GGAAGAAGCA GACCGCTAAC
2101 ACAGTACATA AAAAAGGAGA CATGAACGAT GAACATCAAA AAGTTTGCAA
2151 AACAAGCAAC AGTATTAACC TTTACTACCG CACTGCTGGC AGGAGGCGCA
2201 ACTCAAGCGT TTGCGAAAGA AACGAACCAA AAGCCATATA AGGAAACATA
2251 CGGCATTTCC CATATTACAC GCCATGATAT GCTGCAAATC CCTGAACAGC
2301 AAAAAAATGA AAAATATCAA GTTCCTGAAT TCGATTCGTC CACAATTAAA
2351 AATATCTCTT CTGCAAAAGG CCTGGACGTT TGGGACAGCT GGCCATTACA
2401 AAACGCTGAC GGCACTGTCG CAAACTATCA CGGCTACCAC ATCGTCTTTG
2451 CATTAGCCGG AGATCCTAAA AATGCGGATG ACACATCGAT TTACATGTTC
2501 TATCAAAAAG TCGGCGAAAC TTCTATTGAC AGCTGGAAAA ACGCTGGCCG
2551 CGTCTTTAAA GACAGCGACA AATTCGATGC AAATGATTCT ATCCTAAAAG
2601 ACCAAACACA AGAATGGTCA GGTTCAGCCA CATTTACATC TGACGGAAAA
2651 ATCCGTTTAT TCTACACTGA TTTCTCCGGT AAACATTACG GCAAACAAAC
2701 ACTGACAACT GCACAAGTTA ACGTATCAGC ATCAGACAGC TCTTTGAACA
2751 TCAACGGTGT AGAGGATTAT AAATCAATCT TTGACGGTGA CGGAAAAACG
2801 TATCAAAATG TACAGCAGTT CATCGATGAA GGCAACTACA GCTCAGGCGA
2851 CAACCATACG CTGAGAGATC CTCACTACGT AGAAGATAAA GGCCACAAAT
2901 ACTTAGTATT TGAAGCAAAC ACTGGAACTG AAGATGGCTA CCAAGGCGAA
2951 GAATCTTTAT TTAACAAAGC ATACTATGGC AAAAGCACAT CATTCTTCCG
3001 TCAAGAAAGT CAAAAACTTC TGCAAAGCGA TAAAAAACGC ACGGCTGAGT
3051 TAGCAAACGG CGCTCTCGGT ATGATTGAGC TAAACGATGA TTACACACTG
3101 AAAAAAGTGA TGAAACCGCT GATTGCATCT AACACAGTAA CAGATGAAAT
3151 TGAACGCGCG AACGTCTTTA AAATGAACGG CAAATGGTAC CTGTTCACTG
3201 ACTCCCGCGG ATCAAAAATG ACGATTGACG GCATTACGTC TAACGATATT
3251 TACATGCTTG GTTATGTTTC TAATTCTTTA ACTGGCCCAT ACAAGCCGCT
3301 GAACAAAACT GGCCTTGTGT TAAAAATGGA TCTTGATCCT AACGATGTAA
3351 CCTTTACTTA CTCACACTTC GCTGTACCTC AAGCGAAAGG AAACAATGTC
3401 GTGATTACAA GCTATATGAC AAACAGAGGA TTCTACGCAG ACAAACAATC
3451 AACGTTTGCG CCAAGCTTCC TGCTGAACAT CAAAGGCAAG AAAACATCTG
3501 TTGTCAAAGA CAGCATCCTT GAACAAGGAC AATTAACAGT TAACAAATAA
3551 AAACGCAAAA GAAAATGCCG ATNNCCGGTT TATTGACTAC CGGAAGCAGT
3601 GTGACCGTGT GCTTCTCAAA TGCCTCAGGC TGTCTATGTG TGACTGTTGA
3651 GCTGTAACAA GTTGTCTCAG GTGTTCAATT TCATGTTCTA GTTGCTTTGT
3701 TTTACTGGTT TCACCTGTTC TATTAGGTGT TACATGCTGT TCATCTGTTA
3751 CATTGTCGAT CTGTTCATGG TGAACAGCTT TAAATGCACC AAAAACTCGT
3801 AAAAGCTCTG ATGTATCTAT CTTTTTTACA CCGTTTTCAT CTGTGCATAT
3851 GGACAGTTTT CCCTTTGATA TCTAACGGTG AACAGTTGTT CTACTTTTGT
3901 TTGTTAGTCT TGATGCTTCA CTGATAGATA CAAGAGCCAT AAGAACCTCA
3951 GATCCTTCCG TATTTAGCCA GTATGTTCTC TAGTGTGGTT CGTTGTTTTT
4001 GCGTGAGCCA TGAGAACGAA CCATTGAGAT CATGCTTACT TTGCATGTCA
4051 CTCAAAAATT TTGCCTCAAA ACTGGTGAGC TGAATTTTTG CAGTTAAAGC
4101 ATCGTGTAGT GTTTTTCTTA GTCCGTTACG TAGGTAGGAA TCTGATGTAA
4151 TGGTTGTTGG TATTTTGTCA CCATTCATTT TTATCTGGTT GTTCTCAAGT
4201 TCGGTTACGA GATCCATTTG TCTATCTAGT TCAACTTGGA AAATCAACGT
4251 ATCAGTCGGG CGGCCTCGCT TATCAACCAC CAATTTCATA TTGCTGTAAG
4301 TGTTTAAATC TTTACTTATT GGTTTCAAAA CCCATTGGTT AAGCCTTTTA
4351 AACTCATGGT AGTTATTTTC AAGCATTAAC ATGAACTTAA ATTCATCAAG
4401 GCTAATCTCT ATATTTGCCT TGTGAGTTTT CTTTTGTGTT AGTTCTTTTA
4451 ATAACCACTC ATAAATCCTC ATAGAGTATT TGTTTTCAAA AGACTTAACA
4501 TGTTCCAGAT TATATTTTAT GAATTTTTTT AACTGGAAAA GATAAGGCAA
4551 TATCTCTTCA CTAAAAACTA ATTCTAATTT TTCGCTTGAG AACTTGGCAT
4601 AGTTTGTCCA CTGGAAAATC TCAAAGCCTT TAACCAAAGG ATTCCTGATT
4651 TCCACAGTTC TCGTCATCAG CTCTCTGGTT GCTTTAGCTA ATACACCATA
4701 AGCATTTTCC CTACTGATGT TCATCATCTG AGCGTATTGG TTATAAGTGA
4751 ACGATACCGT CCGTTCTTTC CTTGTAGGGT TTTCAATCGT GGGGTTGAGT
4801 AGTGCCACAC AGCATAAAAT TAGCTTGGTT TCATGCTCCG TTAAGTCATA
4851 GCGACTAATC GCTAGTTCAT TTGCTTTGAA AACAACTAAT TCAGACATAC
4901 ATCTCAATTG GTCTAGGTGA TTTTAATCAC TATACCAATT GAGATGGGCT
4951 AGTCAATGAT AATTACTAGT CCTTTTCCTT TGAGTTGTGG GTATCTGTAA
5001 ATTCTGCTAG ACCTTTGCTG GAAAACTTGT AAATTCTGCT AGACCCTCTG
5051 TAAATTCCGC TAGACCTTTG TGTGTTTTTT TTGTTTATAT TCAAGTGGTT
5101 ATAATTTATA GAATAAAGAA AGAATAAAAA AAGATAAAAA GAATAGATCC
5151 CAGCCCTGTG TATAACTCAC TACTTTAGTC AGTTCCGCAG TATTACAAAA
5201 GGATGTCGCA AACGCTGTTT GCTCCTCTAC AAAACAGACC TTAAAACCCT
5251 AAAGGCTTAA GTAGCACCCT CGCAAGCTCG GGCAAATCGC TGAATATTCC
5301 TTTTGTCTCC GACCATCAGG CACCTGAGTC GCTGTCTTTT TCGTGACATT
5351 CAGTTCGCTG CGCTCACGGC TCTGGCAGTG AATGGGGGTA AATGGCACTA
5401 CAGGCGCCTT TTATGGATTC ATGCAAGGAA ACTACCCATA ATACAAGAAA
5451 AGCCCGTCAC GGGCTTCTCA GGGCGTTTTA TGGCGGGTCT GCTATGTGGT
5501 GCTATCTGAC TTTTTGCTGT TCAGCAGTTC CTGCCCTCTG ATTTTCCAGT
5551 CTGACCACTT CGGATTATCC CGTGACAGGT CATTCAGACT GGCTAATGCA
5601 CCCAGTAAGG CAGCGGTATC ATCAACAGGC TTACCCGTCT TACTGTCnGG
5651 ATCGACGCTC TCCCTTATGC GACTCCTGCA T