LOCUS       Exported                5283 bp DNA     circular SYN 03-SEP-2024
DEFINITION  Bacterial vector with a 6xHis-TEV leader for high-throughput 
            purification of recombinant proteins.
ACCESSION   .
VERSION     .
KEYWORDS    pMCSG7
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5283)
  AUTHORS   Stols L, Gu M, Dieckman L, Raffen R, Collart FR, Donnelly MI.
  TITLE     A new vector for high-throughput, ligation-independent cloning 
            encoding a tobacco etch virus protease cleavage site.
  JOURNAL   Protein Expr. Purif. 2002;25:8-15.
  PUBMED    12071693
REFERENCE   2  (bases 1 to 5283)
  AUTHORS   Midwest Center for Structural Genomics
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 5283)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5283)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Protein
            Expr. Purif."; date: "2002"; volume: "25"; pages: "8-15"
COMMENT     SGRef: number: 2; type: "Journal Article"
COMMENT     SGRef: number: 3; 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..5283
                     /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(268..285)
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             complement(286..288)
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     RBS             complement(296..318)
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    complement(333..357)
                     /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(358..376)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        689..766
                     /label=lacI promoter
     CDS             767..1846
                     /label=lacI
                     /note="lac repressor"
     protein_bind    1862..1883
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             2658..2846
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
     misc_feature    2951..3090
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(3276..3864)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4038..4895)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(4896..4999)
                     /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 ctcggtaccc agatctacac cagaagaatg atgatgatga tggtgcatat gtatatctcc
      301 ttcttaaagt taaacaaaat tatttctaga ggggaattgt tatccgctca caattcccct
      361 atagtgagtc gtattaattt cgcgggatcg agatcgatct cgatcctcta cgccggacgc
      421 atcgtggccg gcatcaccgg cgccacaggt gcggttgctg gcgcctatat cgccgacatc
      481 accgatgggg aagatcgggc tcgccacttc gggctcatga gcgcttgttt cggcgtgggt
      541 atggtggcag gccccgtggc cgggggactg ttgggcgcca tctccttgca tgcaccattc
      601 cttgcggcgg cggtgctcaa cggcctcaac ctactactgg gctgcttcct aatgcaggag
      661 tcgcataagg gagagcgtcg agatcccgga caccatcgaa tggcgcaaaa cctttcgcgg
      721 tatggcatga tagcgcccgg aagagagtca attcagggtg gtgaatgtga aaccagtaac
      781 gttatacgat gtcgcagagt atgccggtgt ctcttatcag accgtttccc gcgtggtgaa
      841 ccaggccagc cacgtttctg cgaaaacgcg ggaaaaagtg gaagcggcga tggcggagct
      901 gaattacatt cccaaccgcg tggcacaaca actggcgggc aaacagtcgt tgctgattgg
      961 cgttgccacc tccagtctgg ccctgcacgc gccgtcgcaa attgtcgcgg cgattaaatc
     1021 tcgcgccgat caactgggtg ccagcgtggt ggtgtcgatg gtagaacgaa gcggcgtcga
     1081 agcctgtaaa gcggcggtgc acaatcttct cgcgcaacgc gtcagtgggc tgatcattaa
     1141 ctatccgctg gatgaccagg atgccattgc tgtggaagct gcctgcacta atgttccggc
     1201 gttatttctt gatgtctctg accagacacc catcaacagt attattttct cccatgaaga
     1261 cggtacgcga ctgggcgtgg agcatctggt cgcattgggt caccagcaaa tcgcgctgtt
     1321 agcgggccca ttaagttctg tctcggcgcg tctgcgtctg gctggctggc ataaatatct
     1381 cactcgcaat caaattcagc cgatagcgga acgggaaggc gactggagtg ccatgtccgg
     1441 ttttcaacaa accatgcaaa tgctgaatga gggcatcgtt cccactgcga tgctggttgc
     1501 caacgatcag atggcgctgg gcgcaatgcg cgccattacc gagtccgggc tgcgcgttgg
     1561 tgcggatatc tcggtagtgg gatacgacga taccgaagac agctcatgtt atatcccgcc
     1621 gttaaccacc atcaaacagg attttcgcct gctggggcaa accagcgtgg accgcttgct
     1681 gcaactctct cagggccagg cggtgaaggg caatcagctg ttgcccgtct cactggtgaa
     1741 aagaaaaacc accctggcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc
     1801 attaatgcag ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa
     1861 ttaatgtaag ttagctcact cattaggcac cgggatctcg accgatgccc ttgagagcct
     1921 tcaacccagt cagctccttc cggtgggcgc ggggcatgac tatcgtcgcc gcacttatga
     1981 ctgtcttctt tatcatgcaa ctcgtaggac aggtgccggc agcgctctgg gtcattttcg
     2041 gcgaggaccg ctttcgctgg agcgcgacga tgatcggcct gtcgcttgcg gtattcggaa
     2101 tcttgcacgc cctcgctcaa gccttcgtca ctggtcccgc caccaaacgt ttcggcgaga
     2161 agcaggccat tatcgccggc atggcggccc cacgggtgcg catgatcgtg ctcctgtcgt
     2221 tgaggacccg gctaggctgg cggggttgcc ttactggtta gcagaatgaa tcaccgatac
     2281 gcgagcgaac gtgaagcgac tgctgctgca aaacgtctgc gacctgagca acaacatgaa
     2341 tggtcttcgg tttccgtgtt tcgtaaagtc tggaaacgcg gaagtcagcg ccctgcacca
     2401 ttatgttccg gatctgcatc gcaggatgct gctggctacc ctgtggaaca cctacatctg
     2461 tattaacgaa gcgctggcat tgaccctgag tgatttttct ctggtcccgc cgcatccata
     2521 ccgccagttg tttaccctca caacgttcca gtaaccgggc atgttcatca tcagtaaccc
     2581 gtatcgtgag catcctctct cgtttcatcg gtatcattac ccccatgaac agaaatcccc
     2641 cttacacgga ggcatcagtg accaaacagg aaaaaaccgc ccttaacatg gcccgcttta
     2701 tcagaagcca gacattaacg cttctggaga aactcaacga gctggacgcg gatgaacagg
     2761 cagacatctg tgaatcgctt cacgaccacg ctgatgagct ttaccgcagc tgcctcgcgc
     2821 gtttcggtga tgacggtgaa aacctctgac acatgcagct cccggagacg gtcacagctt
     2881 gtctgtaagc ggatgccggg agcagacaag cccgtcaggg cgcgtcagcg ggtgttggcg
     2941 ggtgtcgggg cgcagccatg acccagtcac gtagcgatag cggagtgtat actggcttaa
     3001 ctatgcggca tcagagcaga ttgtactgag agtgcaccat tgcggtgtga aataccgcac
     3061 agatgcgtaa ggagaaaata ccgcatcagg cgctcttccg cttcctcgct cactgactcg
     3121 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     3181 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     3241 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     3301 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     3361 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     3421 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     3481 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     3541 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     3601 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     3661 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaaggaca
     3721 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     3781 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     3841 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     3901 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     3961 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     4021 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     4081 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     4141 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     4201 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     4261 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     4321 aatagtttgc gcaacgttgt tgccattgct gcaggcatcg tggtgtcacg ctcgtcgttt
     4381 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     4441 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     4501 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     4561 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     4621 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     4681 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     4741 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     4801 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     4861 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca aattattgaa
     4921 gcatttatca gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata
     4981 aacaaatagg ggttccgcgc acatttcccc gaaaagtgcc acctaaattg taagcgttaa
     5041 tgtgaaccat caccctaatc aagttttttg gggtcgaggt gccgtaaagc actaaatcgg
     5101 aaccctaaag ggagcccccg atttagagct tgacggggaa agccggcgaa cgtggcgaga
     5161 aaggaaggga agaaagcgaa aggagcgggc gctagggcgc tggcaagtgt agcggtcacg
     5221 ctgcgcgtaa ccaccacacc cgccgcgctt aatgcgccgc tacagggcgc gtcccattcg
     5281 cca
//
