pET-52b(+) 3C/LIC vector (V010950)

LOCUS       pET-52b(+)_3C_LI        5227 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Bacterial vector with an HRV 3C site for ligation-independent 
            cloning (LIC) to express proteins tagged with Strep-Tag II.
ACCESSION   .
VERSION     .
KEYWORDS    pET-52b(+) 3C_LIC
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5227)
  AUTHORS   Novagen (EMD Millipore)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 5227)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
COMMENT     For LIC, linearize with SacI and KpnI and treat with T4 DNA 
            polymerase plus dTTP.
FEATURES             Location/Qualifiers
     source          1..5227
                     /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(113..139)
                     /label=9xHis
                     /note="9xHis affinity tag"
     CDS             complement(146..163)
                     /label=thrombin site
                     /note="thrombin recognition and cleavage site"
     CDS             complement(209..232)
                     /label=HRV 3C site
                     /note="recognition and cleavage site for human rhinovirus
                     3C and PreScission proteases"
     CDS             complement(239..262)
                     /label=Strep-Tag II
                     /note="peptide that binds Strep-Tactin(R), an engineered 
                     form
                     of streptavidin"
     CDS             complement(269..271)
                     /codon_start=1
                     /product="start"
                     /label=start
                     /note="ATG"
                     /translation="M"
     RBS             complement(278..300)
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     protein_bind    complement(315..339)
                     /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(340..358)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        671..748
                     /label=lacI promoter
     CDS             749..1828
                     /label=lacI
                     /note="lac repressor"
     protein_bind    1844..1865
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             2403..2591
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
     misc_feature    2696..2838
                     /label=bom
                     /note="basis of mobility region from pBR322"
     rep_origin      complement(3024..3612)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     promoter        3693..3785
                     /label=AmpR promoter
     CDS             3786..4643
                     /label=AmpR
                     /note="beta-lactamase"
     rep_origin      complement(4761..5216)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"