pMCSG46 vector (V010653)

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LOCUS       pMCSG46.        6759 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Bacterial vector with a 6xHN-NusA-TEV leader for high-throughput 
            purification of recombinant proteins.
ACCESSION   .
VERSION     .
KEYWORDS    pMCSG46
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6759)
  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 6759)
  AUTHORS   Midwest Center for Structural Genomics
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 6759)
  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..6759
                     /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(259..1716)
                     /label=NusA
                     /note="transcription elongation protein (N utilization
                     substance protein A)"
     CDS             complement(1717..1752)
                     /label=6xHN
                     /note="6x(His-Asn) affinity tag"
     CDS             complement(1759..1761)
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     RBS             complement(1769..1791)
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    complement(1806..1830)
                     /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(1831..1849)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        2162..2239
                     /label=lacI promoter
     CDS             2240..3319
                     /label=lacI
                     /note="lac repressor"
     protein_bind    3335..3356
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             4131..4319
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
     misc_feature    4424..4566
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(4752..5340)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(5514..6371)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(6372..6475)
                     /label=AmpR promoter