pET32a-LIC vector (V006470)

LOCUS       40924_18471        7889 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pET32a-LIC, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7889)
  AUTHORS   Loppnau P, Kozieradzki I, Arrowsmith C.
  TITLE     Expression vectors for high-throughput In-Fusion cloning
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7889)
  AUTHORS   Loppnau P, Kozieradzki I, Arrowsmith C.
  TITLE     Direct Submission
  JOURNAL   Submitted (19-FEB-2007) Structural Genomics Consortium, University 
            of Toronto, 100 College Street, Toronto, ON M5G1L5, Canada
REFERENCE   3  (bases 1 to 7889)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 7889)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: 
            "Unpublished"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (19-FEB-2007) Structural Genomics Consortium, University of Toronto,
            100 College Street, Toronto, ON M5G1L5, Canada"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..7889
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      12..467
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow
                     indicates direction of (+) strand synthesis"
     promoter        494..598
                     /label=AmpR promoter
     CDS             599..1456
                     /label=AmpR
                     /note="beta-lactamase"
     rep_origin      1630..2218
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     misc_feature    complement(2404..2546)
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(2651..2839)
                     /label=rop
                     /note="Rop protein, which maintains plasmids at low copy
                     number"
     protein_bind    complement(3614..3635)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     CDS             complement(3651..4730)
                     /label=lacI
                     /note="lac repressor"
     promoter        complement(4731..4808)
                     /label=lacI promoter
     promoter        5121..5139
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    5140..5164
                     /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             5179..5201
                     /label=RBS
                     /note="efficient ribosome binding site from bacteriophage
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5209..5535
                     /label=TrxA
                     /note="E. coli thioredoxin"
     CDS             5557..5574
                     /label=6xHis
                     /note="6xHis affinity tag"
     CDS             5584..5601
                     /label=thrombin site
                     /note="thrombin recognition and cleavage site"
     CDS             5608..5652
                     /label=S-Tag
                     /note="affinity and epitope tag derived from pancreatic 
                     ribonuclease A"
     CDS             5668..5688
                     /label=TEV site
                     /note="tobacco etch virus (TEV) protease recognition and 
                     cleavage site"
     protein_bind    complement(5716..5749)
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        5764..6209
                     /label=sacB promoter
                     /note="sacB promoter and control region"
     CDS             6210..7628
                     /label=SacB
                     /note="secreted levansucrase that renders bacterial growth 
                     sensitive to sucrose"
     misc_feature    7704..7718
                     /note="3'-cloning sequence; add complement to downstream
                     PCR primer"
     CDS             7733..7750
                     /label=6xHis
                     /note="6xHis affinity tag"
     terminator      7817..7864
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"