pSecTag2 A

pSecTag2 A

pSecTag2A载体基本信息

pSecTag2A载体质粒图谱和多克隆位点信息

pSecTag2A载体简介

pSecTag2 and pSecTag2/Hygro are mammalian expression vectors designed for the secretion, purification, and detection of fusion proteins.

Each vector has a large multiple cloning site in three reading frames to simplify cloning in frame with the N-terminal secretion signal.

The vectors (Figure 1) offer the following features:

 Secretion signal from the V-J2-C region of the mouse Ig kappa-chain for efficient secretion of recombinant proteins (Figure 2)

 Cytomegalovirus (CMV) promoter for high-level constitutive expression

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

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

 Bovine Growth Hormone (BGH) polyadenylation signal and transcription termination sequence to enhance mRNA stability

 SV40 origin for episomal replication and simple vector rescue in cell lines expressing the large T antigen (e.g., COS-1, COS-7)

The pSecTag2 vectors carry the Zeocin resistance gene for cost-effective selection in mammalian cells. Zeocin selection can also be used in E. coli.

The pSecTag2/Hygro vectors have the Hygromycin B resistance gene for selection of stable mammalian cell lines.

引用及参考文献

Molecular dissection of the alpha-dystroglycan- and integrin-binding sites within the globular domain of human laminin-10.

Ido H, Harada K, Futaki S, Hayashi Y, Nishiuchi R, Natsuka Y, Li S, Wada Y, Combs AC, Ervasti JM, Sekiguchi K, J Biol Chem (2004) 279:10946-10954

Product usage: The 5 cDNA was inserted into the NheI/PmeI sites of pcDNA3.1. For site-directed mutagenesis of the LG3 module, the cDNA fragment encoding LG3 was excised from pcDNA-5LG4–5 with AscI and PmeI and recloned into pSecTag2A at the AscI/PmeI sites. Recombinant protein was produced usi

Prohaptoglobin is proteolytically cleaved in the endoplasmic reticulum by the complement C1r-like protein.Wicher KB, Fries E, Proc Natl Acad Sci U S A (2004) 101:14390-14395

Product usage: HEK293 and COS-1 cells were transiently transfected with C1r-LP cDNA cloned into pcDNA6-myc-his or with proHp cDNA cloned into pSecTag. After 24 h, the cells were rinsed twIce with PBS and grown for 48 h in a serum-free medium. The cell media were then collected.

pSecTag2A载体序列

ORIGIN

    1 GACGGATCGG GAGATCTCCC GATCCCCTAT GGTCGACTCT CAGTACAATC TGCTCTGATG

   61 CCGCATAGTT AAGCCAGTAT CTGCTCCCTG CTTGTGTGTT GGAGGTCGCT GAGTAGTGCG

  121 CGAGCAAAAT TTAAGCTACA ACAAGGCAAG GCTTGACCGA CAATTGCATG AAGAATCTGC

  181 TTAGGGTTAG GCGTTTTGCG CTGCTTCGCG ATGTACGGGC CAGATATACG CGTTGACATT

  241 GATTATTGAC TAGTTATTAA TAGTAATCAA TTACGGGGTC ATTAGTTCAT AGCCCATATA

  301 TGGAGTTCCG CGTTACATAA CTTACGGTAA ATGGCCCGCC TGGCTGACCG CCCAACGACC

  361 CCCGCCCATT GACGTCAATA ATGACGTATG TTCCCATAGT AACGCCAATA GGGACTTTCC

  421 ATTGACGTCA ATGGGTGGAC TATTTACGGT AAACTGCCCA CTTGGCAGTA CATCAAGTGT

  481 ATCATATGCC AAGTACGCCC CCTATTGACG TCAATGACGG TAAATGGCCC GCCTGGCATT

  541 ATGCCCAGTA CATGACCTTA TGGGACTTTC CTACTTGGCA GTACATCTAC GTATTAGTCA

  601 TCGCTATTAC CATGGTGATG CGGTTTTGGC AGTACATCAA TGGGCGTGGA TAGCGGTTTG

  661 ACTCACGGGG ATTTCCAAGT CTCCACCCCA TTGACGTCAA TGGGAGTTTG TTTTGGCACC

  721 AAAATCAACG GGACTTTCCA AAATGTCGTA ACAACTCCGC CCCATTGACG CAAATGGGCG

  781 GTAGGCGTGT ACGGTGGGAG GTCTATATAA GCAGAGCTCT CTGGCTAACT AGAGAACCCA

  841 CTGCTTACTG GCTTATCGAA ATTAATACGA CTCACTATAG GGAGACCCAA GCTGGCTAGC

  901 CACCATGGAG ACAGACACAC TCCTGCTATG GGTACTGCTG CTCTGGGTTC CAGGTTCCAC

  961 TGGTGACGCG GCCCAGCCGG CCAGGCGCGC CGTACGAAGC TTGGTACCGA GCTCGGATCC

 1021 ACTCCAGTGT GGTGGAATTC TGCAGATATC CAGCACAGTG GCGGCCGCTC GAGGAGGGCC

 1081 CGAACAAAAA CTCATCTCAG AAGAGGATCT GAATAGCGCC GTCGACCATC ATCATCATCA

 1141 TCATTGAGTT TAAACCCGCT GATCAGCCTC GACTGTGCCT TCTAGTTGCC AGCCATCTGT

 1201 TGTTTGCCCC TCCCCCGTGC CTTCCTTGAC CCTGGAAGGT GCCACTCCCA CTGTCCTTTC

 1261 CTAATAAAAT GAGGAAATTG CATCGCATTG TCTGAGTAGG TGTCATTCTA TTCTGGGGGG

 1321 TGGGGTGGGG CAGGACAGCA AGGGGGAGGA TTGGGAAGAC AATAGCAGGC ATGCTGGGGA

 1381 TGCGGTGGGC TCTATGGCTT CTGAGGCGGA AAGAACCAGC TGGGGCTCTA GGGGGTATCC

 1441 CCACGCGCCC TGTAGCGGCG CATTAAGCGC GGCGGGTGTG GTGGTTACGC GCAGCGTGAC

 1501 CGCTACACTT GCCAGCGCCC TAGCGCCCGC TCCTTTCGCT TTCTTCCCTT CCTTTCTCGC

 1561 CACGTTCGCC GGCTTTCCCC GTCAAGCTCT AAATCGGGGC ATCCCTTTAG GGTTCCGATT

 1621 TAGTGCTTTA CGGCACCTCG ACCCCAAAAA ACTTGATTAG GGTGATGGTT CACGTAGTGG

 1681 GCCATCGCCC TGATAGACGG TTTTTCGCCC TTTGACGTTG GAGTCCACGT TCTTTAATAG

 1741 TGGACTCTTG TTCCAAACTG GAACAACACT CAACCCTATC TCGGTCTATT CTTTTGATTT

 1801 ATAAGGGATT TTGGGGATTT CGGCCTATTG GTTAAAAAAT GAGCTGATTT AACAAAAATT

 1861 TAACGCGAAT TAATTCTGTG GAATGTGTGT CAGTTAGGGT GTGGAAAGTC CCCAGGCTCC

 1921 CCAGCAGGCA GAAGTATGCA AAGCATGCAT CTCAATTAGT CAGCAACCAG GTGTGGAAAG

 1981 TCCCCAGGCT CCCCAGCAGG CAGAAGTATG CAAAGCATGC ATCTCAATTA GTCAGCAACC

 2041 ATAGTCCCGC CCCTAACTCC GCCCATCCCG CCCCTAACTC CGCCCAGTTC CGCCCATTCT

 2101 CCGCCCCATG GCTGACTAAT TTTTTTTATT TATGCAGAGG CCGAGGCCGC CTCTGCCTCT

 2161 GAGCTATTCC AGAAGTAGTG AGGAGGCTTT TTTGGAGGCC TAGGCTTTTG CAAAAAGCTC

 2221 CCGGGAGCTT GTATATCCAT TTTCGGATCT GATCAGCACG TGTTGACAAT TAATCATCGG

 2281 CATAGTATAT CGGCATAGTA TAATACGACA AGGTGAGGAA CTAAACCATG GCCAAGTTGA

 2341 CCAGTGCCGT TCCGGTGCTC ACCGCGCGCG ACGTCGCCGG AGCGGTCGAG TTCTGGACCG

 2401 ACCGGCTCGG GTTCTCCCGG GACTTCGTGG AGGACGACTT CGCCGGTGTG GTCCGGGACG

 2461 ACGTGACCCT GTTCATCAGC GCGGTCCAGG ACCAGGTGGT GCCGGACAAC ACCCTGGCCT

 2521 GGGTGTGGGT GCGCGGCCTG GACGAGCTGT ACGCCGAGTG GTCGGAGGTC GTGTCCACGA

 2581 ACTTCCGGGA CGCCTCCGGG CCGGCCATGA CCGAGATCGG CGAGCAGCCG TGGGGGCGGG

 2641 AGTTCGCCCT GCGCGACCCG GCCGGCAACT GCGTGCACTT CGTGGCCGAG GAGCAGGACT

 2701 GACACGTGCT ACGAGATTTC GATTCCACCG CCGCCTTCTA TGAAAGGTTG GGCTTCGGAA

 2761 TCGTTTTCCG GGACGCCGGC TGGATGATCC TCCAGCGCGG GGATCTCATG CTGGAGTTCT

 2821 TCGCCCACCC CAACTTGTTT ATTGCAGCTT ATAATGGTTA CAAATAAAGC AATAGCATCA

 2881 CAAATTTCAC AAATAAAGCA TTTTTTTCAC TGCATTCTAG TTGTGGTTTG TCCAAACTCA

 2941 TCAATGTATC TTATCATGTC TGTATACCGT CGACCTCTAG CTAGAGCTTG GCGTAATCAT

 3001 GGTCATAGCT GTTTCCTGTG TGAAATTGTT ATCCGCTCAC AATTCCACAC AACATACGAG

 3061 CCGGAAGCAT AAAGTGTAAA GCCTGGGGTG CCTAATGAGT GAGCTAACTC ACATTAATTG

 3121 CGTTGCGCTC ACTGCCCGCT TTCCAGTCGG GAAACCTGTC GTGCCAGCTG CATTAATGAA

 3181 TCGGCCAACG CGCGGGGAGA GGCGGTTTGC GTATTGGGCG CTCTTCCGCT TCCTCGCTCA

 3241 CTGACTCGCT GCGCTCGGTC GTTCGGCTGC GGCGAGCGGT ATCAGCTCAC TCAAAGGCGG

 3301 TAATACGGTT ATCCACAGAA TCAGGGGATA ACGCAGGAAA GAACATGTGA GCAAAAGGCC

 3361 AGCAAAAGGC CAGGAACCGT AAAAAGGCCG CGTTGCTGGC GTTTTTCCAT AGGCTCCGCC

 3421 CCCCTGACGA GCATCACAAA AATCGACGCT CAAGTCAGAG GTGGCGAAAC CCGACAGGAC

 3481 TATAAAGATA CCAGGCGTTT CCCCCTGGAA GCTCCCTCGT GCGCTCTCCT GTTCCGACCC

 3541 TGCCGCTTAC CGGATACCTG TCCGCCTTTC TCCCTTCGGG AAGCGTGGCG CTTTCTCAAT

 3601 GCTCACGCTG TAGGTATCTC AGTTCGGTGT AGGTCGTTCG CTCCAAGCTG GGCTGTGTGC

 3661 ACGAACCCCC CGTTCAGCCC GACCGCTGCG CCTTATCCGG TAACTATCGT CTTGAGTCCA

 3721 ACCCGGTAAG ACACGACTTA TCGCCACTGG CAGCAGCCAC TGGTAACAGG ATTAGCAGAG

 3781 CGAGGTATGT AGGCGGTGCT ACAGAGTTCT TGAAGTGGTG GCCTAACTAC GGCTACACTA

 3841 GAAGGACAGT ATTTGGTATC TGCGCTCTGC TGAAGCCAGT TACCTTCGGA AAAAGAGTTG

 3901 GTAGCTCTTG ATCCGGCAAA CAAACCACCG CTGGTAGCGG TGGTTTTTTT GTTTGCAAGC

 3961 AGCAGATTAC GCGCAGAAAA AAAGGATCTC AAGAAGATCC TTTGATCTTT TCTACGGGGT

 4021 CTGACGCTCA GTGGAACGAA AACTCACGTT AAGGGATTTT GGTCATGAGA TTATCAAAAA

 4081 GGATCTTCAC CTAGATCCTT TTAAATTAAA AATGAAGTTT TAAATCAATC TAAAGTATAT

 4141 ATGAGTAAAC TTGGTCTGAC AGTTACCAAT GCTTAATCAG TGAGGCACCT ATCTCAGCGA

 4201 TCTGTCTATT TCGTTCATCC ATAGTTGCCT GACTCCCCGT CGTGTAGATA ACTACGATAC

 4261 GGGAGGGCTT ACCATCTGGC CCCAGTGCTG CAATGATACC GCGAGACCCA CGCTCACCGG

 4321 CTCCAGATTT ATCAGCAATA AACCAGCCAG CCGGAAGGGC CGAGCGCAGA AGTGGTCCTG

 4381 CAACTTTATC CGCCTCCATC CAGTCTATTA ATTGTTGCCG GGAAGCTAGA GTAAGTAGTT

 4441 CGCCAGTTAA TAGTTTGCGC AACGTTGTTG CCATTGCTAC AGGCATCGTG GTGTCACGCT

 4501 CGTCGTTTGG TATGGCTTCA TTCAGCTCCG GTTCCCAACG ATCAAGGCGA GTTACATGAT

 4561 CCCCCATGTT GTGCAAAAAA GCGGTTAGCT CCTTCGGTCC TCCGATCGTT GTCAGAAGTA

 4621 AGTTGGCCGC AGTGTTATCA CTCATGGTTA TGGCAGCACT GCATAATTCT CTTACTGTCA

 4681 TGCCATCCGT AAGATGCTTT TCTGTGACTG GTGAGTACTC AACCAAGTCA TTCTGAGAAT

 4741 AGTGTATGCG GCGACCGAGT TGCTCTTGCC CGGCGTCAAT ACGGGATAAT ACCGCGCCAC

 4801 ATAGCAGAAC TTTAAAAGTG CTCATCATTG GAAAACGTTC TTCGGGGCGA AAACTCTCAA

 4861 GGATCTTACC GCTGTTGAGA TCCAGTTCGA TGTAACCCAC TCGTGCACCC AACTGATCTT

 4921 CAGCATCTTT TACTTTCACC AGCGTTTCTG GGTGAGCAAA AACAGGAAGG CAAAATGCCG

 4981 CAAAAAAGGG AATAAGGGCG ACACGGAAAT GTTGAATACT CATACTCTTC CTTTTTCAAT

 5041 ATTATTGAAG CATTTATCAG GGTTATTGTC TCATGAGCGG ATACATATTT GAATGTATTT

 5101 AGAAAAATAA ACAAATAGGG GTTCCGCGCA CATTTCCCCG AAAAGTGCCA CCTGACGTC

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