pkk223-3

pkk223-3

pkk223-3载体基本信息

pkk223-3载体质粒图谱和多克隆位点信息

pkk223-3载体简介

pKK223-3 (1) contains the strong tac promoter upstream from themultiple cloning site and the strong rrnB ribosomal terminator downstreamfor control of protein expression (2-4).

Induction: tac promoter is inducible with 1-5 mM IPTG. Promoter isvery strong; even uninduced cells may show a low level of expression.

Expression: Genes containing a ribosome-binding site and ATG can beinserted into any unique site in the MCS for expression. The ribosomebindingsite on the plasmid can be utilised to express inserts if the ATGstart codon of the insert is within 5-13 bp from the provided ribosomebindingsite (5, 6).

Transcription terminators: rrnB transcription terminators stabilize theplasmid by inhibiting read-through transcription initiated from the tacpromoter in the parent plasmid.

Host(s): lac Iq strains; E. coli JM105 is recommended.

Selectable marker(s): Plasmid confers resistance to 100 μg/ml ampicillin.

Amplification: Recommended.

Control Regions: Expression control region: tac promoter: -10: 50-44; -35: 72-67; Ribosome binding site: 11. MCS: 4552-4587. rrnB operonregion: 5S rRNA region: 4471-4352; rrnB T1 terminator: 4348-4305;rrnB T2 terminator: 4173-4146. b-lactamase gene region: Promoter: -10:4082-4077; -35: 4105-4100; Start codon (ATG): 4035; Stop codon(TAA): 3178. Plasmid replication region: Site of replication initiation:2417-2415; Region necessary for replication: 3111-2415.

pkk223-3载体序列

ORIGIN

    1 TTCTGTTTCC TGTGTGAAAT TGTTATCCGC TCACAATTCC ACACATTATA CGAGCCGATG

   61 ATTAATTGTC AACAGCTCAT TTCAGAATAT TTGCCAGAAC CGTTATGATG TCGGCGCAAA

  121 AAACATTATC CAGAACGGGA GTGCGCCTTG AGCGACACGA ATTATGCAGT GATTTACGAC

  181 CTGCACAGCC ATACCACAGC TTCCGATGGC TGCCTGACGC CAGAAGCATT GGTGCACCGT

  241 GCAGTCGATA AGCTCCGGAT CCTCTACGCC GGACGCATCG TGGCCGGCAT CACCGGCGCC

  301 ACAGGTGCGG TTGCTGGCGC CTATATCGCC GACATCACCG ATGGGGAAGA TCGGGCTCGC

  361 CACTTCGGGC TCATGAGCGC TTGTTTCGGC GTGGGTATGG TGGCAGGCCC CGTGGCCGGG

  421 GGACTGTTGG GCGCCATCTC CTTGCATGCA CCATTCCTTG CGGCGGCGGT GCTCAACGGC

  481 CTCAACCTAC TACTGGGCTG CTTCCTAATG CAGGAGTCGC ATAAGGGAGA GCGTCGACCG

  541 ATGCCCTTGA GAGCCTTCAA CCCAGTCAGC TCCTTCCGGT GGGCGCGGGG CATGACTATC

  601 GTCGCCGCAC TTATGACTGT CTTCTTTATC ATGCAACTCG TAGGACAGGT GCCGGCAGCG

  661 CTCTGGGTCA TTTTCGGCGA GGACCGCTTT CGCTGGAGCG CGACGATGAT CGGCCTGTCG

  721 CTTGCGGTAT TCGGAATCTT GCACGCCCTC GCTCAAGCCT TCGTCACTGG TCCCGCCACC

  781 AAACGTTTCG GCGAGAAGCA GGCCATTATC GCCGGCATGG CGGCCGACGC GCTGGGCTAC

  841 GTCTTGCTGG CGTTCGCGAC GCGAGGCTGG ATGGCCTTCC CCATTATGAT TCTTCTCGCT

  901 TCCGGCGGCA TCGGGATGCC CGCGTTGCAG GCCATGCTGT CCAGGCAGGT AGATGACGAC

  961 CATCAGGGAC AGCTTCAAGG ATCGCTCGCG GCTCTTACCA GCCTAACTTC GATCACTGGA

 1021 CCGCTGATCG TCACGGCGAT TTATGCCGCC TCGGCGAGCA CATGGAACGG GTTGGCATGG

 1081 ATTGTAGGCG CCGCCCTATA CCTTGTCTGC CTCCCCGCGT TGCGTCGCGG TGCATGGAGC

 1141 CGGGCCACCT CGACCTGAAT GGAAGCCGGC GGCACCTCGC TAACGGATTC ACCACTCCAA

 1201 GAATTGGAGC CAATCAATTC TTGCGGAGAA CTGTGAATGC GCAAACCAAC CCTTGGCAGA

 1261 ACATATCCAT CGCGTCCGCC ATCTCCAGCA GCCGCACGCG GCGCATCTCG GGCAGCGTTG

 1321 GGTCCTGGCC ACGGGTGCGC ATGATCGTGC TCCTGTCGTT GAGGACCCGG CTAGGCTGGC

 1381 GGGGTTGCCT TACTGGTTAG CAGAATGAAT CACCGATACG CGAGCGAACG TGAAGCGACT

 1441 GCTGCTGCAA AACGTCTGCG ACCTGAGCAA CAACATGAAT GGTCTTCGGT TTCCGTGTTT

 1501 CGTAAAGTCT GGAAACGCGG AAGTCAGCGC CCTGCACCAT TATGTTCCGG ATCTGCATCG

 1561 CAGGATGCTG CTGGCTACCC TGTGGAACAC CTACATCTGT ATTAACGAAG CGCTGGCATT

 1621 GACCCTGAGT GATTTTTCTC TGGTCCCGCC GCATCCATAC CGCCAGTTGT TTACCCTCAC

 1681 AACGTTCCAG TAACCGGGCA TGTTCATCAT CAGTAACCCG TATCGTGAGC ATCCTCTCTC

 1741 GTTTCATCGG TATCATTACC CCCATGAACA GAAATTCCCC CTTACACGGA GGCATCAAGT

 1801 GACCAAACAG GAAAAAACCG CCCTTAACAT GGCCCGCTTT ATCAGAAGCC AGACATTAAC

 1861 GCTTCTGGAG AAACTCAACG AGCTGGACGC GGATGAACAG GCAGACATCT GTGAATCGCT

 1921 TCACGACCAC GCTGATGAGC TTTACCGCAG CTGCCTCGCG CGTTTCGGTG ATGACGGTGA

 1981 AAACCTCTGA CACATGCAGC TCCCGGAGAC GGTCACAGCT TGTCTGTAAG CGGATGCCGG

 2041 GAGCAGACAA GCCCGTCAGG GCGCGTCAGC GGGTGTTGGC GGGTGTCGGG GCGCAGCCAT

 2101 GACCCAGTCA CGTAGCGATA GCGGAGTGTA TACTGGCTTA ACTATGCGGC ATCAGAGCAG

 2161 ATTGTACTGA GAGTGCACCA TATGCGGTGT GAAATACCGC ACAGATGCGT AAGGAGAAAA

 2221 TACCGCATCA GGCGCTCTTC CGCTTCCTCG CTCACTGACT CGCTGCGCTC GGTCGTTCGG

 2281 CTGCGGCGAG CGGTATCAGC TCACTCAAAG GCGGTAATAC GGTTATCCAC AGAATCAGGG

 2341 GATAACGCAG GAAAGAACAT GTGAGCAAAA GGCCAGCAAA AGGCCAGGAA CCGTAAAAAG

 2401 GCCGCGTTGC TGGCGTTTTT CCATAGGCTC CGCCCCCCTG ACGAGCATCA CAAAAATCGA

 2461 CGCTCAAGTC AGAGGTGGCG AAACCCGACA GGACTATAAA GATACCAGGC GTTTCCCCCT

 2521 GGAAGCTCCC TCGTGCGCTC TCCTGTTCCG ACCCTGCCGC TTACCGGATA CCTGTCCGCC

 2581 TTTCTCCCTT CGGGAAGCGT GGCGCTTTCT CATAGCTCAC GCTGTAGGTA TCTCAGTTCG

 2641 GTGTAGGTCG TTCGCTCCAA GCTGGGCTGT GTGCACGAAC CCCCCGTTCA GCCCGACCGC

 2701 TGCGCCTTAT CCGGTAACTA TCGTCTTGAG TCCAACCCGG TAAGACACGA CTTATCGCCA

 2761 CTGGCAGCAG CCACTGGTAA CAGGATTAGC AGAGCGAGGT ATGTAGGCGG TGCTACAGAG

 2821 TTCTTGAAGT GGTGGCCTAA CTACGGCTAC ACTAGAAGGA CAGTATTTGG TATCTGCGCT

 2881 CTGCTGAAGC CAGTTACCTT CGGAAAAAGA GTTGGTAGCT CTTGATCCGG CAAACAAACC

 2941 ACCGCTGGTA GCGGTGGTTT TTTTGTTTGC AAGCAGCAGA TTACGCGCAG AAAAAAAGGA

 3001 TCTCAAGAAG ATCCTTTGAT CTTTTCTACG GGGTCTGACG CTCAGTGGAA CGAAAACTCA

 3061 CGTTAAGGGA TTTTGGTCAT GAGATTATCA AAAAGGATCT TCACCTAGAT CCTTTTAAAT

 3121 TAAAAATGAA GTTTTAAATC AATCTAAAGT ATATATGAGT AAACTTGGTC TGACAGTTAC

 3181 CAATGCTTAA TCAGTGAGGC ACCTATCTCA GCGATCTGTC TATTTCGTTC ATCCATAGTT

 3241 GCCTGACTCC CCGTCGTGTA GATAACTACG ATACGGGAGG GCTTACCATC TGGCCCCAGT

 3301 GCTGCAATGA TACCGCGAGA CCCACGCTCA CCGGCTCCAG ATTTATCAGC AATAAACCAG

 3361 CCAGCCGGAA GGGCCGAGCG CAGAAGTGGT CCTGCAACTT TATCCGCCTC CATCCAGTCT

 3421 ATTAATTGTT GCCGGGAAGC TAGAGTAAGT AGTTCGCCAG TTAATAGTTT GCGCAACGTT

 3481 GTTGCCATTG CTACAGGCAT CGTGGTGTCA CGCTCGTCGT TTGGTATGGC TTCATTCAGC

 3541 TCCGGTTCCC AACGATCAAG GCGAGTTACA TGATCCCCCA TGTTGTGCAA AAAAGCGGTT

 3601 AGCTCCTTCG GTCCTCCGAT CGTTGTCAGA AGTAAGTTGG CCGCAGTGTT ATCACTCATG

 3661 GTTATGGCAG CACTGCATAA TTCTCTTACT GTCATGCCAT CCGTAAGATG CTTTTCTGTG

 3721 ACTGGTGAGT ACTCAACCAA GTCATTCTGA GAATAGTGTA TGCGGCGACC GAGTTGCTCT

 3781 TGCCCGGCGT CAACACGGGA TAATACCGCG CCACATAGCA GAACTTTAAA AGTGCTCATC

 3841 ATTGGAAAAC GTTCTTCGGG GCGAAAACTC TCAAGGATCT TACCGCTGTT GAGATCCAGT

 3901 TCGATGTAAC CCACTCGTGC ACCCAACTGA TCTTCAGCAT CTTTTACTTT CACCAGCGTT

 3961 TCTGGGTGAG CAAAAACAGG AAGGCAAAAT GCCGCAAAAA AGGGAATAAG GGCGACACGG

 4021 AAATGTTGAA TACTCATACT CTTCCTTTTT CAATATTATT GAAGCATTTA TCAGGGTTAT

 4081 TGTCTCATGA GCGGATACAT ATTTGAATGT ATTTAGAAAA ATAAACAAAA GAGTTTGTAG

 4141 AAACGCAAAA AGGCCATCCG TCAGGATGGC CTTCTGCTTA ATTTGATGCC TGGCAGTTTA

 4201 TGGCGGGCGT CCTGCCCGCC ACCCTCCGGG CCGTTGCTTC GCAACGTTCA AATCCGCTCC

 4261 CGGCGGATTT GTCCTACTCA GGAGAGCGTT CACCGACAAA CAACAGATAA AACGAAAGGC

 4321 CCAGTCTTTC GACTGAGCCT TTCGTTTTAT TTGATGCCTG GCAGTTCCCT ACTCTCGCAT

 4381 GGGGAGACCC CACACTACCA TCGGCGCTAC GGCGTTTCAC TTCTGAGTTC GGCATGGGGT

 4441 CAGGTGGGAC CACCGCGCTA CTGCCGCCAG GCAAATTCTG TTTTATCAGA CCGCTTCTGC

 4501 GTTCTGATTT AATCTGTATC AGGCTGAAAA TCTTCTCTCA TCCGCCAAAA CAGAAGCTTG

 4561 GCTGCAGGTC GACGGATCCC CGGGAA

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