LOCUS       pMCSG11.        4079 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Bacterial coexpression vector with a 6xHis-TEV leader and a p15A 
            origin, for high-throughput purification of recombinant proteins.
ACCESSION   .
VERSION     .
KEYWORDS    pMCSG11
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4079)
  AUTHORS   Eschenfeldt WH, Lucy S, Millard CS, Joachimiak A, Mark ID.
  TITLE     A family of LIC vectors for high-throughput cloning and purification
            of proteins.
  JOURNAL   Methods Mol. Biol. 2009;498:105-15.
  PUBMED    18988021
REFERENCE   2  (bases 1 to 4079)
  AUTHORS   Midwest Center for Structural Genomics
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 4079)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Methods 
            Mol. Biol."; date: "2009"; volume: "498"; pages: "105-15"
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..4079
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    3..27
                     /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)."
     RBS             42..64
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             71..73
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     CDS             83..100
                     /label=6xHis
                     /note="6xHis affinity tag"
     promoter        214..232
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    233..257
                     /label=lac repressor encoded by lacI binding site
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="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)."
     RBS             286..291
                     /note="ribosome binding site"
     CDS             300..302
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     CDS             303..320
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             345..365
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
     CDS             437..481
                     /label=S-Tag
                     /note="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
     terminator      533..580
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     CDS             complement(803..1459)
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
     promoter        complement(1460..1562)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding
                     chloramphenicol acetyltransferase"
     rep_origin      complement(2088..2632)
                     /direction=LEFT
                     /label=p15A ori
                     /note="Plasmids containing the medium-copy-number p15A
                     origin of replication can be propagated in E. coli cells 
                     that contain a second plasmid with the ColE1 origin."
     protein_bind    complement(2823..2844)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(2860..3939)
                     /label=lacI
                     /note="lac repressor"
     promoter        complement(3940..4017)
                     /label=lacI promoter
ORIGIN
        1 ggggaattgt gagcggataa caattcccct gtagaaataa ttttgtttaa ctttaataag
       61 gagatatacc atgggcagca gccatcacca tcatcaccac agccaggatc cgaattcgag
      121 ctcggcgcgc ctgcaggtcg acaagcttgc ggccgcataa tgcttaagtc gaacagaaag
      181 taatcgtatt gtacacggcc gcataatcga aattaatacg actcactata ggggaattgt
      241 gagcggataa caattcccca tcttagtata ttagttaagt ataagaagga gatatacata
      301 tgcaccatca tcatcatcat tcttctggtg tagatctggg taccgagaac ctgtacttcc
      361 aatccaatat tggaagtgga taacggatcc gaattcgagc tccgtcgaca agcttgcggc
      421 cgcactcgag tctggtaaag aaaccgctgc tgcgaaattt gaacgccagc acatggactc
      481 gtctactagc gcagcttaat taacctaggc tgctgccacc gctgagcaat aactagcata
      541 accccttggg gcctctaaac gggtcttgag gggttttttg ctgaaacctc aggcatttga
      601 gaagcacacg gtcacactgc ttccggtagt caataaaccg gtaaaccagc aatagacata
      661 agcggctatt taacgaccct gccctgaacc gacgaccggg tcgaatttgc tttcgaattt
      721 ctgccattca tccgcttatt atcacttatt caggcgtagc accaggcgtt taagggcacc
      781 aataactgcc ttaaaaaaat tacgccccgc cctgccactc atcgcagtac tgttgtaatt
      841 cattaagcat tctgccgaca tggaagccat cacagacggc atgatgaacc tgaatcgcca
      901 gcggcatcag caccttgtcg ccttgcgtat agtatttgcc catagtgaaa acgggggcga
      961 agaagttgtc catattggcc acgtttaaat caaaactggt gaaactcacc cagggattgg
     1021 ctgagacgaa aaacatattc tcaataaacc ctttagggaa ataggccagg ttttcaccgt
     1081 aacacgccac atcttgcgaa tatatgtgta gaaactgccg gaaatcgtcg tggtattcac
     1141 tccagagcga tgaaaacgtt tcagtttgct catggaaaac ggtgtaacaa gggtgaacac
     1201 tatcccatat caccagctca ccgtctttca ttgccatacg gaactccgga tgagcattca
     1261 tcaggcgggc aagaatgtga ataaaggccg gataaaactt gtgcttattt ttctttacgg
     1321 tctttaaaaa ggccgtaata tccagctgaa cggtctggtt ataggtacat tgagcaactg
     1381 actgaaatgc ctcaaaatgt tctttacgat gccattggga tatatcaacg gtggtatatc
     1441 cagtgatttt tttctccatt ttagcttcct tagctcctga aaatctcgat aactcaaaaa
     1501 atacgcccgg tagtgatctt atttcattat ggtgaaagtt ggaacctctt acgtgccgat
     1561 caacgtctca ttttcgccaa aagttggccc agggcttccc ggtatcaaca gggacaccag
     1621 gatttattta ttctgcgaag tgatcttccg tcacaggtat ttattcggcg caaagtgcgt
     1681 cgggtgatgc tgccaactta ctgatttagt gtatgatggt gtttttgagg tgctccagtg
     1741 gcttctgttt ctatcagctg tccctcctgt tcagctactg acggggtggt gcgtaacggc
     1801 aaaagcaccg ccggacatca gcgctagcgg agtgtatact ggcttactat gttggcactg
     1861 atgagggtgt cagtgaagtg cttcatgtgg caggagaaaa aaggctgcac cggtgcgtca
     1921 gcagaatatg tgatacagga tatattccgc ttcctcgctc actgactcgc tacgctcggt
     1981 cgttcgactg cggcgagcgg aaatggctta cgaacggggc ggagatttcc tggaagatgc
     2041 caggaagata cttaacaggg aagtgagagg gccgcggcaa agccgttttt ccataggctc
     2101 cgcccccctg acaagcatca cgaaatctga cgctcaaatc agtggtggcg aaacccgaca
     2161 ggactataaa gataccaggc gtttcccctg gcggctccct cgtgcgctct cctgttcctg
     2221 cctttcggtt taccggtgtc attccgctgt tatggccgcg tttgtctcat tccacgcctg
     2281 acactcagtt ccgggtaggc agttcgctcc aagctggact gtatgcacga accccccgtt
     2341 cagtccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggaaagacat
     2401 gcaaaagcac cactggcagc agccactggt aattgattta gaggagttag tcttgaagtc
     2461 atgcgccggt taaggctaaa ctgaaaggac aagttttggt gactgcgctc ctccaagcca
     2521 gttacctcgg ttcaaagagt tggtagctca gagaaccttc gaaaaaccgc cctgcaaggc
     2581 ggttttttcg ttttcagagc aagagattac gcgcagacca aaacgatctc aagaagatca
     2641 tcttattaat cagataaatt atttctagat ttcagtgcaa tttatctctt caaatgtagc
     2701 acctgaagtc agccccatac gatataagtt gtaattctca tgttagtcat gccccgcgcc
     2761 caccggaagg agctgactgg gttgaaggct ctcaagggca tcggtcgaga tcccggtgcc
     2821 taatgagtga gctaacttac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga
     2881 aacctgtcgt gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt
     2941 attgggcgcc agggtggttt ttcttttcac cagtgagacg ggcaacagct gattgccctt
     3001 caccgcctgg ccctgagaga gttgcagcaa gcggtccacg ctggtttgcc ccagcaggcg
     3061 aaaatcctgt ttgatggtgg ttaacggcgg gatataacat gagctgtctt cggtatcgtc
     3121 gtatcccact accgagatgt ccgcaccaac gcgcagcccg gactcggtaa tggcgcgcat
     3181 tgcgcccagc gccatctgat cgttggcaac cagcatcgca gtgggaacga tgccctcatt
     3241 cagcatttgc atggtttgtt gaaaaccgga catggcactc cagtcgcctt cccgttccgc
     3301 tatcggctga atttgattgc gagtgagata tttatgccag ccagccagac gcagacgcgc
     3361 cgagacagaa cttaatgggc ccgctaacag cgcgatttgc tggtgaccca atgcgaccag
     3421 atgctccacg cccagtcgcg taccgtcttc atgggagaaa ataatactgt tgatgggtgt
     3481 ctggtcagag acatcaagaa ataacgccgg aacattagtg caggcagctt ccacagcaat
     3541 ggcatcctgg tcatccagcg gatagttaat gatcagccca ctgacgcgtt gcgcgagaag
     3601 attgtgcacc gccgctttac aggcttcgac gccgcttcgt tctaccatcg acaccaccac
     3661 gctggcaccc agttgatcgg cgcgagattt aatcgccgcg acaatttgcg acggcgcgtg
     3721 cagggccaga ctggaggtgg caacgccaat cagcaacgac tgtttgcccg ccagttgttg
     3781 tgccacgcgg ttgggaatgt aattcagctc cgccatcgcc gcttccactt tttcccgcgt
     3841 tttcgcagaa acgtggctgg cctggttcac cacgcgggaa acggtctgat aagagacacc
     3901 ggcatactct gcgacatcgt ataacgttac tggtttcaca ttcaccaccc tgaattgact
     3961 ctcttccggg cgctatcatg ccataccgcg aaaggttttg cgccattcga tggtgtccgg
     4021 gatctcgacg ctctccctta tgcgactcct gcattaggaa attaatacga ctcactata
//
