pOPING-ET vector (V004229)

LOCUS       40924_33777        5588 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pOPING-ET, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5588)
  AUTHORS   Nettleship JE, Ren J, Rahman N, Berrow NS, Hatherley D, Barclay AN, 
            Owens RJ.
  TITLE     A pipeline for the production of antibody fragments for structural 
            studies using transient expression in HEK 293T cells
  JOURNAL   Protein Expr. Purif. 62 (1), 83-89 (2008)
  PUBMED    18662785
REFERENCE   2  (bases 1 to 5588)
  AUTHORS   Berrow NS, Owens RJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (20-MAY-2008) Oxford Protein Production Facility, Oxford 
            University, Roosevelt Drive, Oxford, Oxfordshire OX3 7BN, UK
REFERENCE   3  (bases 1 to 5588)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5588)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Protein 
            Expr. Purif."; date: "2008"; volume: "62"; issue: "1"; pages: 
            "83-89"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (20-MAY-2008) Oxford Protein Production Facility, Oxford University,
            Roosevelt Drive, Oxford, Oxfordshire OX3 7BN, UK"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5588
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_recomb     181..1008
                     /label=baculovirus recombination region (lef2/ORF603)
                     /note="contains ORF603 and part of lef2"
     enhancer        1140..1443
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1449..1726
                     /label=chicken beta-actin promoter
     promoter        2150..2168
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    2169..2193
                     /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        2209..2318
                     /label=p10 promoter
                     /note="baculovirus promoter for expression in insect cells"
     misc_feature    2333..2416
                     /label=mu-phosphatase secretion leader peptide
                     /note="mu-phosphatase secretion leader peptide"
     misc_feature    2403..2417
                     /label=5' infusion site after KpnI digest
                     /note="5' infusion site after KpnI digest"
     protein_bind    2463..2484
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     promoter        2499..2529
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    2537..2553
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /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)."
     CDS             2573..2746
                     /codon_start=1
                     /label=lacZ-alpha
                     /note="LacZ-alpha fragment of beta-galactosidase"
                     /translation="MTMITDSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEART
                     DRPSQQLRSLNGE"
     CDS             2776..2793
                     /codon_start=1
                     /label=6xHis
                     /note="6xHis affinity tag"
                     /translation="HHHHHH"
     polyA_signal    2948..3003
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     terminator      3091..3138
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     misc_recomb     3151..3856
                     /label=baculovirus recombination region (ORF1629)
                     /note="contains part of ORF1629"
     protein_bind    complement(3865..3898)
                     /label=loxP
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      complement(4066..4653)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4669..5526)
                     /codon_start=1
                     /label=AmpR
                     /note="beta-lactamase"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"