pSpeedET vector (V010574)

LOCUS       pSpeedET.        5464 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Vector with a lethal ccdB gene, for polymerase incomplete primer 
            extension (PIPE) cloning to express proteins with a cleavable 6xHis 
            tag.
ACCESSION   .
VERSION     .
KEYWORDS    pSpeedET
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5464)
  AUTHORS   Klock HE, Koesema EJ, Knuth MW, Lesley SA.
  TITLE     Combining the polymerase incomplete primer extension method for 
            cloning and mutagenesis with microscreening to accelerate structural
            genomics efforts.
  JOURNAL   Proteins 2008;71:982-94.
  PUBMED    18004753
REFERENCE   2  (bases 1 to 5464)
  AUTHORS   Joint Center for Structural Genomics (JCSG)
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 5464)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Proteins"; 
            date: "2008"; volume: "71"; pages: "982-94"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5464
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        1..285
                     /label=araBAD promoter
                     /note="promoter of the L-arabinose operon of E. coli; the
                     araC regulatory gene is transcribed in the opposite 
                     direction (Guzman et al., 1995)"
     promoter        291..309
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     RBS             312..334
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             346..348
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     CDS             364..381
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             382..402
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
     promoter        418..448
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an 'up' mutation"
     CDS             502..1158
                     /label=CmR
                     /note="chloramphenicol acetyltransferase"
     CDS             1503..1805
                     /label=ccdB
                     /note="CcdB, a bacterial toxin that poisons DNA gyrase"
     terminator      2079..2165
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     terminator      2257..2284
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB
                     gene"
     promoter        2304..2395
                     /label=AmpR promoter
     CDS             2396..3208
                     /label=KanR
                     /note="aminoglycoside phosphotransferase"
     rep_origin      3382..3970
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    complement(4156..4296)
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(4563..5438)
                     /label=araC
                     /note="L-arabinose regulatory protein"