pBad/Myc-His C

pBad/Myc-His C

pBadMyc-His C载体基本信息

pBadMyc-His C载体质粒图谱和多克隆位点信息

pBadMyc-His 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/Myc-His A,B,C 载体简介

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.

The pBAD/Myc-His Kit provides all of the necessary reagents to express your protein in a tightly regulated fashion. The pBAD/Myc-His vector expresses native proteins or fusion proteins with a C-terminal tag. The vector provides:

Þ      The araBAD promoter for tightly regulated expression

Þ      Translation initiation signals optimized for E. coliexpression

Þ      C-terminal polyhistidine (6xHis) tag for purification with nickel-chelating resin or detection with an Anti-His(C-term) Antibody

Þ      C-terminal c-myc epitope for detection and analysis with an Anti-myc Antibody

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

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.

pBadMyc-His 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 GGATCCGAGC TCGAGATCTG CAGCTGGTAC CATATGGGAA

  361 TTCGAAGCTT ACGTAGAACA AAAACTCATC TCAGAAGAGG ATCTGAATAG CGCCGTCGAC

  421 CATCATCATC ATCATCATTG AGTTTAAACG GTCTCCAGCT TGGCTGTTTT GGCGGATGAG

  481 AGAAGATTTT CAGCCTGATA CAGATTAAAT CAGAACGCAG AAGCGGTCTG ATAAAACAGA

  541 ATTTGCCTGG CGGCAGTAGC GCGGTGGTCC CACCTGACCC CATGCCGAAC TCAGAAGTGA

  601 AACGCCGTAG CGCCGATGGT AGTGTGGGGT CTCCCCATGC GAGAGTAGGG AACTGCCAGG

  661 CATCAAATAA AACGAAAGGC TCAGTCGAAA GACTGGGCCT TTCGTTTTAT CTGTTGTTTG

  721 TCGGTGAACG CTCTCCTGAG TAGGACAAAT CCGCCGGGAG CGGATTTGAA CGTTGCGAAG

  781 CAACGGCCCG GAGGGTGGCG GGCAGGACGC CCGCCATAAA CTGCCAGGCA TCAAATTAAG

  841 CAGAAGGCCA TCCTGACGGA TGGCCTTTTT GCGTTTCTAC AAACTCTTTT GTTTATTTTT

  901 CTAAATACAT TCAAATATGT ATCCGCTCAT GAGACAATAA CCCTGATAAA TGCTTCAATA

  961 ATATTGAAAA AGGAAGAGTA TGAGTATTCA ACATTTCCGT GTCGCCCTTA TTCCCTTTTT

 1021 TGCGGCATTT TGCCTTCCTG TTTTTGCTCA CCCAGAAACG CTGGTGAAAG TAAAAGATGC

 1081 TGAAGATCAG TTGGGTGCAC GAGTGGGTTA CATCGAACTG GATCTCAACA GCGGTAAGAT

 1141 CCTTGAGAGT TTTCGCCCCG AAGAACGTTT TCCAATGATG AGCACTTTTA AAGTTCTGCT

 1201 ATGTGGCGCG GTATTATCCC GTGTTGACGC CGGGCAAGAG CAACTCGGTC GCCGCATACA

 1261 CTATTCTCAG AATGACTTGG TTGAGTACTC ACCAGTCACA GAAAAGCATC TTACGGATGG

 1321 CATGACAGTA AGAGAATTAT GCAGTGCTGC CATAACCATG AGTGATAACA CTGCGGCCAA

 1381 CTTACTTCTG ACAACGATCG GAGGACCGAA GGAGCTAACC GCTTTTTTGC ACAACATGGG

 1441 GGATCATGTA ACTCGCCTTG ATCGTTGGGA ACCGGAGCTG AATGAAGCCA TACCAAACGA

 1501 CGAGCGTGAC ACCACGATGC CTGTAGCAAT GGCAACAACG TTGCGCAAAC TATTAACTGG

 1561 CGAACTACTT ACTCTAGCTT CCCGGCAACA ATTAATAGAC TGGATGGAGG CGGATAAAGT

 1621 TGCAGGACCA CTTCTGCGCT CGGCCCTTCC GGCTGGCTGG TTTATTGCTG ATAAATCTGG

 1681 AGCCGGTGAG CGTGGGTCTC GCGGTATCAT TGCAGCACTG GGGCCAGATG GTAAGCCCTC

 1741 CCGTATCGTA GTTATCTACA CGACGGGGAG TCAGGCAACT ATGGATGAAC GAAATAGACA

 1801 GATCGCTGAG ATAGGTGCCT CACTGATTAA GCATTGGTAA CTGTCAGACC AAGTTTACTC

 1861 ATATATACTT TAGATTGATT TAAAACTTCA TTTTTAATTT AAAAGGATCT AGGTGAAGAT

 1921 CCTTTTTGAT AATCTCATGA CCAAAATCCC TTAACGTGAG TTTTCGTTCC ACTGAGCGTC

 1981 AGACCCCGTA GAAAAGATCA AAGGATCTTC TTGAGATCCT TTTTTTCTGC GCGTAATCTG

 2041 CTGCTTGCAA ACAAAAAAAC CACCGCTACC AGCGGTGGTT TGTTTGCCGG ATCAAGAGCT

 2101 ACCAACTCTT TTTCCGAAGG TAACTGGCTT CAGCAGAGCG CAGATACCAA ATACTGTCCT

 2161 TCTAGTGTAG CCGTAGTTAG GCCACCACTT CAAGAACTCT GTAGCACCGC CTACATACCT

 2221 CGCTCTGCTA ATCCTGTTAC CAGTGGCTGC TGCCAGTGGC GATAAGTCGT GTCTTACCGG

 2281 GTTGGACTCA AGACGATAGT TACCGGATAA GGCGCAGCGG TCGGGCTGAA CGGGGGGTTC

 2341 GTGCACACAG CCCAGCTTGG AGCGAACGAC CTACACCGAA CTGAGATACC TACAGCGTGA

 2401 GCTATGAGAA AGCGCCACGC TTCCCGAAGG GAGAAAGGCG GACAGGTATC CGGTAAGCGG

 2461 CAGGGTCGGA ACAGGAGAGC GCACGAGGGA GCTTCCAGGG GGAAACGCCT GGTATCTTTA

 2521 TAGTCCTGTC GGGTTTCGCC ACCTCTGACT TGAGCGTCGA TTTTTGTGAT GCTCGTCAGG

 2581 GGGGCGGAGC CTATGGAAAA ACGCCAGCAA CGCGGCCTTT TTACGGTTCC TGGCCTTTTG

 2641 CTGGCCTTTT GCTCACATGT TCTTTCCTGC GTTATCCCCT GATTCTGTGG ATAACCGTAT

 2701 TACCGCCTTT GAGTGAGCTG ATACCGCTCG CCGCAGCCGA ACGACCGAGC GCAGCGAGTC

 2761 AGTGAGCGAG GAAGCGGAAG AGCGCCTGAT GCGGTATTTT CTCCTTACGC ATCTGTGCGG

 2821 TATTTCACAC CGCATATGGT GCACTCTCAG TACAATCTGC TCTGATGCCG CATAGTTAAG

 2881 CCAGTATACA CTCCGCTATC GCTACGTGAC TGGGTCATGG CTGCGCCCCG ACACCCGCCA

 2941 ACACCCGCTG ACGCGCCCTG ACGGGCTTGT CTGCTCCCGG CATCCGCTTA CAGACAAGCT

 3001 GTGACCGTCT CCGGGAGCTG CATGTGTCAG AGGTTTTCAC CGTCATCACC GAAACGCGCG

 3061 AGGCAGCAGA TCAATTCGCG CGCGAAGGCG AAGCGGCATG CATAATGTGC CTGTCAAATG

 3121 GACGAAGCAG GGATTCTGCA AACCCTATGC TACTCCGTCA AGCCGTCAAT TGTCTGATTC

 3181 GTTACCAATT ATGACAACTT GACGGCTACA TCATTCACTT TTTCTTCACA ACCGGCACGG

 3241 AACTCGCTCG GGCTGGCCCC GGTGCATTTT TTAAATACCC GCGAGAAATA GAGTTGATCG

 3301 TCAAAACCAA CATTGCGACC GACGGTGGCG ATAGGCATCC GGGTGGTGCT CAAAAGCAGC

 3361 TTCGCCTGGC TGATACGTTG GTCCTCGCGC CAGCTTAAGA CGCTAATCCC TAACTGCTGG

 3421 CGGAAAAGAT GTGACAGACG CGACGGCGAC AAGCAAACAT GCTGTGCGAC GCTGGCGATA

 3481 TCAAAATTGC TGTCTGCCAG GTGATCGCTG ATGTACTGAC AAGCCTCGCG TACCCGATTA

 3541 TCCATCGGTG GATGGAGCGA CTCGTTAATC GCTTCCATGC GCCGCAGTAA CAATTGCTCA

 3601 AGCAGATTTA TCGCCAGCAG CTCCGAATAG CGCCCTTCCC CTTGCCCGGC GTTAATGATT

 3661 TGCCCAAACA GGTCGCTGAA ATGCGGCTGG TGCGCTTCAT CCGGGCGAAA GAACCCCGTA

 3721 TTGGCAAATA TTGACGGCCA GTTAAGCCAT TCATGCCAGT AGGCGCGCGG ACGAAAGTAA

 3781 ACCCACTGGT GATACCATTC GCGAGCCTCC GGATGACGAC CGTAGTGATG AATCTCTCCT

 3841 GGCGGGAACA GCAAAATATC ACCCGGTCGG CAAACAAATT CTCGTCCCTG ATTTTTCACC

 3901 ACCCCCTGAC CGCGAATGGT GAGATTGAGA ATATAACCTT TCATTCCCAG CGGTCGGTCG

 3961 ATAAAAAAAT CGAGATAACC GTTGGCCTCA ATCGGCGTTA AACCCGCCAC CAGATGGGCA

 4021 TTAAACGAGT ATCCCGGCAG CAGGGGATCA TTTTGCGCTT CAGCCATACT TTTCATACTC

 4081 CCGCCATTCA GAG

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