pBad/His C

pBad/His C

pBadHis C载体基本信息

pBadHis C载体质粒图谱和多克隆位点信息

pBadHis C载体简介

pBAD/His和PBAD/Myc-His载体质粒是衍生于pBR322载体。载体设计用来在大肠杆菌中进行可调节，剂量依赖性的表达和纯化重组目的蛋白。使用大肠杆菌araBAD启动子（pBAD）增强了大肠杆菌重组蛋白可溶性表达的水平。pBAD/His和pBAD/Myc His载体上的调节蛋白AraC能够调控pBad启动子。

pBAD/His A,B,C 载体介绍

The pBAD/His Kit provides all of the necessary reagents to express your protein in a tightly regulated fashion. The vector pBAD/His allows you to express your protein with an N-terminal tag. The vector provides:

Þ      The araBAD promoter for tightly regulated expression

Þ      Translation initiation signals optimized for E. coliexpression

Þ      N-terminal polyhistidine (6xHis) tag for purification with nickel-chelating resin and detection with an Anti-

HisG Antibody

Þ      N-terminal Xpress epitope for detection and analysis with an Anti-Xpress Antibody

Þ      Enterokinase cleavage site for removing the N-terminal tag following purification

Three vectors are provided (A, B, and C). Each has the N-terminal tag in a different reading frame relative to the multiple cloning site to simplify in-frame cloning of your gene.

The pBAD/His and pBAD/Myc-His plasmids are pBR322-derived expression vectors designed for regulated, dose-dependent recombinant protein expression and purification in E. coli. Optimum levels of soluble, recombinant protein are possible using the araBAD promoter (PBAD) from E. coli. The regulatory protein, AraC, is provided on the pBAD/His and pBAD/Myc-His vectors allowing regulation of PBAD.

L-阿拉伯糖调控表达

In the presence of L-arabinose, expression from PBAD is turned on while the absence of L-arabinose produces very low levels of transcription from PBAD (Lee, 1980; Lee et al., 1987). Uninduced levels are repressed even further by growth in the presence of glucose. Glucose reduces the levels of 3′,5′-cyclic AMP, thus lowering expression of the catabolite-repressed PBAD promoter (Miyada et al., 1984). By varying the concentration of L-arabinose, protein expression levels can be optimized to ensure maximum expression of soluble protein. In addition, the tight regulation of PBAD by AraC is useful for expression of potentially toxic or essential genes (Carson et al., 1991; Dalbey and Wickner, 1985; Guzman et al., 1992; Kuhn and Wickner, 1985; Russell et al., 1989; San Millan et al., 1989). For more information on the mechanism of expression and repression of the ara regulon, refer to Schleif, 1992.

pBadHis C载体序列

ORIGIN

    1 AAGAAACCAA TTGTCCATAT TGCATCAGAC ATTGCCGTCA CTGCGTCTTT TACTGGCTCT

   61 TCTCGCTAAC CAAACCGGTA ACCCCGCTTA TTAAAAGCAT TCTGTAACAA AGCGGGACCA

  121 AAGCCATGAC AAAAACGCGT AACAAAAGTG TCTATAATCA CGGCAGAAAA GTCCACATTG

  181 ATTATTTGCA CGGCGTCACA CTTTGCTATG CCATAGCATT TTTATCCATA AGATTAGCGG

  241 ATCCTACCTG ACGCTTTTTA TCGCAACTCT CTACTGTTTC TCCATACCCG TTTTTTGGGC

  301 TAACAGGAGG AATTAACCAT GGGGGGTTCT CATCATCATC ATCATCATGG TATGGCTAGC

  361 ATGACTGGTG GACAGCAAAT GGGTCGGGAT CTGTACGACG ATGACGATAA GGATCGATGG

  421 ATCCGACCTC GAGATCTGCA GATGGTACCA TATGGGAATT CGAAGCTTGG CTGTTTTGGC

  481 GGATGAGAGA AGATTTTCAG CCTGATACAG ATTAAATCAG AACGCAGAAG CGGTCTGATA

  541 AAACAGAATT TGCCTGGCGG CAGTAGCGCG GTGGTCCCAC CTGACCCCAT GCCGAACTCA

  601 GAAGTGAAAC GCCGTAGCGC CGATGGTAGT GTGGGGTCTC CCCATGCGAG AGTAGGGAAC

  661 TGCCAGGCAT CAAATAAAAC GAAAGGCTCA GTCGAAAGAC TGGGCCTTTC GTTTTATCTG

  721 TTGTTTGTCG GTGAACGCTC TCCTGAGTAG GACAAATCCG CCGGGAGCGG ATTTGAACGT

  781 TGCGAAGCAA CGGCCCGGAG GGTGGCGGGC AGGACGCCCG CCATAAACTG CCAGGCATCA

  841 AATTAAGCAG AAGGCCATCC TGACGGATGG CCTTTTTGCG TTTCTACAAA CTCTTTTGTT

  901 TATTTTTCTA AATACATTCA AATATGTATC CGCTCATGAG ACAATAACCC TGATAAATGC

  961 TTCAATAATA TTGAAAAAGG AAGAGTATGA GTATTCAACA TTTCCGTGTC GCCCTTATTC

 1021 CCTTTTTTGC GGCATTTTGC CTTCCTGTTT TTGCTCACCC AGAAACGCTG GTGAAAGTAA

 1081 AAGATGCTGA AGATCAGTTG GGTGCACGAG TGGGTTACAT CGAACTGGAT CTCAACAGCG

 1141 GTAAGATCCT TGAGAGTTTT CGCCCCGAAG AACGTTTTCC AATGATGAGC ACTTTTAAAG

 1201 TTCTGCTATG TGGCGCGGTA TTATCCCGTG TTGACGCCGG GCAAGAGCAA CTCGGTCGCC

 1261 GCATACACTA TTCTCAGAAT GACTTGGTTG AGTACTCACC AGTCACAGAA AAGCATCTTA

 1321 CGGATGGCAT GACAGTAAGA GAATTATGCA GTGCTGCCAT AACCATGAGT GATAACACTG

 1381 CGGCCAACTT ACTTCTGACA ACGATCGGAG GACCGAAGGA GCTAACCGCT TTTTTGCACA

 1441 ACATGGGGGA TCATGTAACT CGCCTTGATC GTTGGGAACC GGAGCTGAAT GAAGCCATAC

 1501 CAAACGACGA GCGTGACACC ACGATGCCTG TAGCAATGGC AACAACGTTG CGCAAACTAT

 1561 TAACTGGCGA ACTACTTACT CTAGCTTCCC GGCAACAATT AATAGACTGG ATGGAGGCGG

 1621 ATAAAGTTGC AGGACCACTT CTGCGCTCGG CCCTTCCGGC TGGCTGGTTT ATTGCTGATA

 1681 AATCTGGAGC CGGTGAGCGT GGGTCTCGCG GTATCATTGC AGCACTGGGG CCAGATGGTA

 1741 AGCCCTCCCG TATCGTAGTT ATCTACACGA CGGGGAGTCA GGCAACTATG GATGAACGAA

 1801 ATAGACAGAT CGCTGAGATA GGTGCCTCAC TGATTAAGCA TTGGTAACTG TCAGACCAAG

 1861 TTTACTCATA TATACTTTAG ATTGATTTAA AACTTCATTT TTAATTTAAA AGGATCTAGG

 1921 TGAAGATCCT TTTTGATAAT CTCATGACCA AAATCCCTTA ACGTGAGTTT TCGTTCCACT

 1981 GAGCGTCAGA CCCCGTAGAA AAGATCAAAG GATCTTCTTG AGATCCTTTT TTTCTGCGCG

 2041 TAATCTGCTG CTTGCAAACA AAAAAACCAC CGCTACCAGC GGTGGTTTGT TTGCCGGATC

 2101 AAGAGCTACC AACTCTTTTT CCGAAGGTAA CTGGCTTCAG CAGAGCGCAG ATACCAAATA

 2161 CTGTCCTTCT AGTGTAGCCG TAGTTAGGCC ACCACTTCAA GAACTCTGTA GCACCGCCTA

 2221 CATACCTCGC TCTGCTAATC CTGTTACCAG TGGCTGCTGC CAGTGGCGAT AAGTCGTGTC

 2281 TTACCGGGTT GGACTCAAGA CGATAGTTAC CGGATAAGGC GCAGCGGTCG GGCTGAACGG

 2341 GGGGTTCGTG CACACAGCCC AGCTTGGAGC GAACGACCTA CACCGAACTG AGATACCTAC

 2401 AGCGTGAGCT ATGAGAAAGC GCCACGCTTC CCGAAGGGAG AAAGGCGGAC AGGTATCCGG

 2461 TAAGCGGCAG GGTCGGAACA GGAGAGCGCA CGAGGGAGCT TCCAGGGGGA AACGCCTGGT

 2521 ATCTTTATAG TCCTGTCGGG TTTCGCCACC TCTGACTTGA GCGTCGATTT TTGTGATGCT

 2581 CGTCAGGGGG GCGGAGCCTA TGGAAAAACG CCAGCAACGC GGCCTTTTTA CGGTTCCTGG

 2641 CCTTTTGCTG GCCTTTTGCT CACATGTTCT TTCCTGCGTT ATCCCCTGAT TCTGTGGATA

 2701 ACCGTATTAC CGCCTTTGAG TGAGCTGATA CCGCTCGCCG CAGCCGAACG ACCGAGCGCA

 2761 GCGAGTCAGT GAGCGAGGAA GCGGAAGAGC GCCTGATGCG GTATTTTCTC CTTACGCATC

 2821 TGTGCGGTAT TTCACACCGC ATATGGTGCA CTCTCAGTAC AATCTGCTCT GATGCCGCAT

 2881 AGTTAAGCCA GTATACACTC CGCTATCGCT ACGTGACTGG GTCATGGCTG CGCCCCGACA

 2941 CCCGCCAACA CCCGCTGACG CGCCCTGACG GGCTTGTCTG CTCCCGGCAT CCGCTTACAG

 3001 ACAAGCTGTG ACCGTCTCCG GGAGCTGCAT GTGTCAGAGG TTTTCACCGT CATCACCGAA

 3061 ACGCGCGAGG CAGCAGATCA ATTCGCGCGC GAAGGCGAAG CGGCATGCAT AATGTGCCTG

 3121 TCAAATGGAC GAAGCAGGGA TTCTGCAAAC CCTATGCTAC TCCGTCAAGC CGTCAATTGT

 3181 CTGATTCGTT ACCAATTATG ACAACTTGAC GGCTACATCA TTCACTTTTT CTTCACAACC

 3241 GGCACGGAAC TCGCTCGGGC TGGCCCCGGT GCATTTTTTA AATACCCGCG AGAAATAGAG

 3301 TTGATCGTCA AAACCAACAT TGCGACCGAC GGTGGCGATA GGCATCCGGG TGGTGCTCAA

 3361 AAGCAGCTTC GCCTGGCTGA TACGTTGGTC CTCGCGCCAG CTTAAGACGC TAATCCCTAA

 3421 CTGCTGGCGG AAAAGATGTG ACAGACGCGA CGGCGACAAG CAAACATGCT GTGCGACGCT

 3481 GGCGATATCA AAATTGCTGT CTGCCAGGTG ATCGCTGATG TACTGACAAG CCTCGCGTAC

 3541 CCGATTATCC ATCGGTGGAT GGAGCGACTC GTTAATCGCT TCCATGCGCC GCAGTAACAA

 3601 TTGCTCAAGC AGATTTATCG CCAGCAGCTC CGAATAGCGC CCTTCCCCTT GCCCGGCGTT

 3661 AATGATTTGC CCAAACAGGT CGCTGAAATG CGGCTGGTGC GCTTCATCCG GGCGAAAGAA

 3721 CCCCGTATTG GCAAATATTG ACGGCCAGTT AAGCCATTCA TGCCAGTAGG CGCGCGGACG

 3781 AAAGTAAACC CACTGGTGAT ACCATTCGCG AGCCTCCGGA TGACGACCGT AGTGATGAAT

 3841 CTCTCCTGGC GGGAACAGCA AAATATCACC CGGTCGGCAA ACAAATTCTC GTCCCTGATT

 3901 TTTCACCACC CCCTGACCGC GAATGGTGAG ATTGAGAATA TAACCTTTCA TTCCCAGCGG

 3961 TCGGTCGATA AAAAAATCGA GATAACCGTT GGCCTCAATC GGCGTTAAAC CCGCCACCAG

 4021 ATGGGCATTA AACGAGTATC CCGGCAGCAG GGGATCATTT TGCGCTTCAG CCATACTTTT

 4081 CATACTCCCG CCATTCAGAG

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pBad/His C其他大肠杆菌表达载体：