LOCUS       pMCSG37.        6426 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Bacterial vector with a 10xHis-MBP-6xHis-TEV leader for 
            high-throughput purification of recombinant proteins.
ACCESSION   .
VERSION     .
KEYWORDS    pMCSG37
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6426)
  AUTHORS   Donnelly MI, Zhou M, Millard CS, Clancy S, Stols L, Eschenfeldt WH, 
            Collart FR, Joachimiak A.
  TITLE     An expression vector tailored for large-scale, high-throughput 
            purification of recombinant proteins.
  JOURNAL   Protein Expr. Purif. 2006;47:446-54.
  PUBMED    16497515
REFERENCE   2  (bases 1 to 6426)
  AUTHORS   Midwest Center for Structural Genomics
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 6426)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Protein 
            Expr. Purif."; date: "2006"; volume: "47"; pages: "446-54"
COMMENT     SGRef: number: 2; type: "Journal Article"
COMMENT     For ligation-independent cloning (LIC), linearize with SspI and 
            treat with T4 DNA polymerase plus dGTP.
FEATURES             Location/Qualifiers
     source          1..6426
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     terminator      complement(26..73)
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(140..157)
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             complement(223..243)
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
     CDS             complement(250..267)
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             complement(274..1371)
                     /label=MBP
                     /note="maltose binding protein from E. coli"
     CDS             complement(1399..1425)
                     /label=9xHis
                     /note="9xHis affinity tag"
     CDS             complement(1426..1428)
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     RBS             complement(1436..1458)
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    complement(1473..1497)
                     /label=lac operator
                     /note="The lac repressor binds to the lac operator to
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(1498..1516)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        1829..1906
                     /label=lacI promoter
     CDS             1907..2986
                     /label=lacI
                     /note="lac repressor"
     protein_bind    3002..3023
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             3798..3986
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
     misc_feature    4091..4233
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(4419..5007)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5181..6038)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(6039..6142)
                     /label=AmpR promoter
ORIGIN
        1 atccggatat agttcctcct ttcagcaaaa aacccctcaa gacccgttta gaggccccaa
       61 ggggttatgc tagttattgc tcagcggtgg cagcagccaa ctcagcttcc tttcgggctt
      121 tgttagcagc cggatctcag tggtggtggt ggtggtgctc gagtgcggcc gcaagcttgt
      181 cgacggagct cgaattcgga tccgttatcc acttccaata ttggattgga agtacaggtt
      241 ctcggtacag tggtggtggt ggtggtgcga attagtctgc gcgtctttca gggcttcatc
      301 gacagtctga cgaccgctgg cggcgttgat caccgcagta cgcacggcat accagaaagc
      361 ggacatctgc gggatgttcg gcatgatttc acctttctgg gcgttttcca tggtggcggc
      421 aatacgtgga tctttcgcca actcttcctc gtaagacttc agcgctacgg cacccagcgg
      481 tttgtcttta ttaaccgctt ccagaccttc atcagtcagc agatagtttt cgaggaactc
      541 ttttgccagc tctttgttcg gactggcggc gttaatacct gcgctcagca cgccaacgaa
      601 cggtttggat ggttgaccct tgaaggtcgg cagtaccgtt acaccataat tcactttgct
      661 ggtgtcgatg ttggaccatg cccacgggcc gttgatggtc atcgctgttt cgcctttatt
      721 aaaggcagct tctgcgatgg agtaatcggt gtctgcattc atgtgtttgt ttttaatcag
      781 gtcaaccagg aaggtcagac ccgctttcgc gccagcgtta tccacgccca cgtctttaat
      841 gtcgtacttg ccgttttcat acttgaacgc ataacccccg tcagcagcaa tcagcggcca
      901 ggtgaagtac ggttcttgca ggttgaacat cagcgcgctc ttacctttcg ctttcagttc
      961 tttatccagc gccgggatct cttcccaggt ttttggcggg ttcggcagca gatctttgtt
     1021 ataaatcagc gataacgctt caacagcgat cgggtaagca atcagcttgc cgttgtaacg
     1081 tacggcatcc caggtaaacg gatacagctt gtcctggaac gctttgtccg gggtgatttc
     1141 agccaacagg ccagattgag cgtagccacc aaagcggtcg tgtgcccaga agataatgtc
     1201 agggccatcg ccagttgccg caacctgtgg gaatttctct tccagtttat ccggatgctc
     1261 aacggtgact ttaattccgg tatctttctc gaatttctta ccgacttcag cgagaccgtt
     1321 atagccttta tcgccgttaa tccagattac cagtttacct tcttcgattt tggtacccag
     1381 atctacacca gaagagtggt ggtggtgatg atgatgatga tggtgcatat gtatatctcc
     1441 ttcttaaagt taaacaaaat tatttctaga ggggaattgt tatccgctca caattcccct
     1501 atagtgagtc gtattaattt cgcgggatcg agatcgatct cgatcctcta cgccggacgc
     1561 atcgtggccg gcatcaccgg cgccacaggt gcggttgctg gcgcctatat cgccgacatc
     1621 accgatgggg aagatcgggc tcgccacttc gggctcatga gcgcttgttt cggcgtgggt
     1681 atggtggcag gccccgtggc cgggggactg ttgggcgcca tctccttgca tgcaccattc
     1741 cttgcggcgg cggtgctcaa cggcctcaac ctactactgg gctgcttcct aatgcaggag
     1801 tcgcataagg gagagcgtcg agatcccgga caccatcgaa tggcgcaaaa cctttcgcgg
     1861 tatggcatga tagcgcccgg aagagagtca attcagggtg gtgaatgtga aaccagtaac
     1921 gttatacgat gtcgcagagt atgccggtgt ctcttatcag accgtttccc gcgtggtgaa
     1981 ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct
     2041 gaattacatt cccaaccgcg tggcacaaca actggcgggc aaacagtcgt tgctgattgg
     2101 cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc
     2161 tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga
     2221 agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa
     2281 ctatccgctg gatgaccagg atgccattgc tgtggaagct gcctgcacta atgttccggc
     2341 gttatttctt gatgtctctg accagacacc catcaacagt attattttct cccatgaaga
     2401 cggtacgcga ctgggcgtgg agcatctggt cgcattgggt caccagcaaa tcgcgctgtt
     2461 agcgggccca ttaagttctg tctcggcgcg tctgcgtctg gctggctggc ataaatatct
     2521 cactcgcaat caaattcagc cgatagcgga acgggaaggc gactggagtg ccatgtccgg
     2581 ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt cccactgcga tgctggttgc
     2641 caacgatcag atggcgctgg gcgcaatgcg cgccattacc gagtccgggc tgcgcgttgg
     2701 tgcggatatc tcggtagtgg gatacgacga taccgaagac agctcatgtt atatcccgcc
     2761 gttaaccacc atcaaacagg attttcgcct gctggggcaa accagcgtgg accgcttgct
     2821 gcaactctct cagggccagg cggtgaaggg caatcagctg ttgcccgtct cactggtgaa
     2881 aagaaaaacc accctggcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
     2941 attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa
     3001 ttaatgtaag ttagctcact cattaggcac cgggatctcg accgatgccc ttgagagcct
     3061 tcaacccagt cagctccttc cggtgggcgc ggggcatgac tatcgtcgcc gcacttatga
     3121 ctgtcttctt tatcatgcaa ctcgtaggac aggtgccggc agcgctctgg gtcattttcg
     3181 gcgaggaccg ctttcgctgg agcgcgacga tgatcggcct gtcgcttgcg gtattcggaa
     3241 tcttgcacgc cctcgctcaa gccttcgtca ctggtcccgc caccaaacgt ttcggcgaga
     3301 agcaggccat tatcgccggc atggcggccc cacgggtgcg catgatcgtg ctcctgtcgt
     3361 tgaggacccg gctaggctgg cggggttgcc ttactggtta gcagaatgaa tcaccgatac
     3421 gcgagcgaac gtgaagcgac tgctgctgca aaacgtctgc gacctgagca acaacatgaa
     3481 tggtcttcgg tttccgtgtt tcgtaaagtc tggaaacgcg gaagtcagcg ccctgcacca
     3541 ttatgttccg gatctgcatc gcaggatgct gctggctacc ctgtggaaca cctacatctg
     3601 tattaacgaa gcgctggcat tgaccctgag tgatttttct ctggtcccgc cgcatccata
     3661 ccgccagttg tttaccctca caacgttcca gtaaccgggc atgttcatca tcagtaaccc
     3721 gtatcgtgag catcctctct cgtttcatcg gtatcattac ccccatgaac agaaatcccc
     3781 cttacacgga ggcatcagtg accaaacagg aaaaaaccgc ccttaacatg gcccgcttta
     3841 tcagaagcca gacattaacg cttctggaga aactcaacga gctggacgcg gatgaacagg
     3901 cagacatctg tgaatcgctt cacgaccacg ctgatgagct ttaccgcagc tgcctcgcgc
     3961 gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt
     4021 gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg
     4081 ggtgtcgggg cgcagccatg acccagtcac gtagcgatag cggagtgtat actggcttaa
     4141 ctatgcggca tcagagcaga ttgtactgag agtgcaccat atatgcggtg tgaaataccg
     4201 cacagatgcg taaggagaaa ataccgcatc aggcgctctt ccgcttcctc gctcactgac
     4261 tcgctgcgct cggtcgttcg gctgcggcga gcggtatcag ctcactcaaa ggcggtaata
     4321 cggttatcca cagaatcagg ggataacgca ggaaagaaca tgtgagcaaa aggccagcaa
     4381 aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt tccataggct ccgcccccct
     4441 gacgagcatc acaaaaatcg acgctcaagt cagaggtggc gaaacccgac aggactataa
     4501 agataccagg cgtttccccc tggaagctcc ctcgtgcgct ctcctgttcc gaccctgccg
     4561 cttaccggat acctgtccgc ctttctccct tcgggaagcg tggcgctttc tcatagctca
     4621 cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca agctgggctg tgtgcacgaa
     4681 ccccccgttc agcccgaccg ctgcgcctta tccggtaact atcgtcttga gtccaacccg
     4741 gtaagacacg acttatcgcc actggcagca gccactggta acaggattag cagagcgagg
     4801 tatgtaggcg gtgctacaga gttcttgaag tggtggccta actacggcta cactagaagg
     4861 acagtatttg gtatctgcgc tctgctgaag ccagttacct tcggaaaaag agttggtagc
     4921 tcttgatccg gcaaacaaac caccgctggt agcggtggtt tttttgtttg caagcagcag
     4981 attacgcgca gaaaaaaagg atctcaagaa gatcctttga tcttttctac ggggtctgac
     5041 gctcagtgga acgaaaactc acgttaaggg attttggtca tgagattatc aaaaaggatc
     5101 ttcacctaga tccttttaaa ttaaaaatga agttttaaat caatctaaag tatatatgag
     5161 taaacttggt ctgacagtta ccaatgctta atcagtgagg cacctatctc agcgatctgt
     5221 ctatttcgtt catccatagt tgcctgactc cccgtcgtgt agataactac gatacgggag
     5281 ggcttaccat ctggccccag tgctgcaatg ataccgcgag acccacgctc accggctcca
     5341 gatttatcag caataaacca gccagccgga agggccgagc gcagaagtgg tcctgcaact
     5401 ttatccgcct ccatccagtc tattaattgt tgccgggaag ctagagtaag tagttcgcca
     5461 gttaatagtt tgcgcaacgt tgttgccatt gctgcaggca tcgtggtgtc acgctcgtcg
     5521 tttggtatgg cttcattcag ctccggttcc caacgatcaa ggcgagttac atgatccccc
     5581 atgttgtgca aaaaagcggt tagctccttc ggtcctccga tcgttgtcag aagtaagttg
     5641 gccgcagtgt tatcactcat ggttatggca gcactgcata attctcttac tgtcatgcca
     5701 tccgtaagat gcttttctgt gactggtgag tactcaacca agtcattctg agaatagtgt
     5761 atgcggcgac cgagttgctc ttgcccggcg tcaatacggg ataataccgc gccacatagc
     5821 agaactttaa aagtgctcat cattggaaaa cgttcttcgg ggcgaaaact ctcaaggatc
     5881 ttaccgctgt tgagatccag ttcgatgtaa cccactcgtg cacccaactg atcttcagca
     5941 tcttttactt tcaccagcgt ttctgggtga gcaaaaacag gaaggcaaaa tgccgcaaaa
     6001 aagggaataa gggcgacacg gaaatgttga atactcatac tcttcctttt tcaaattatt
     6061 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     6121 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctaaa ttgtaagcgt
     6181 taatgtgaac catcacccta atcaagtttt ttggggtcga ggtgccgtaa agcactaaat
     6241 cggaacccta aagggagccc ccgatttaga gcttgacggg gaaagccggc gaacgtggcg
     6301 agaaaggaag ggaagaaagc gaaaggagcg ggcgctaggg cgctggcaag tgtagcggtc
     6361 acgctgcgcg taaccaccac acccgccgcg cttaatgcgc cgctacaggg cgcgtcccat
     6421 tcgcca
//
