Basic Vector Information
- Vector Name:
- pOPING-ET
- Antibiotic Resistance:
- Ampicillin
- Length:
- 5588 bp
- Type:
- Expression vector
- Replication origin:
- ori
- Source/Author:
- Nettleship JE, Ren J, Rahman N, Berrow NS, Hatherley D, Barclay AN, Owens RJ.
- Promoter:
- chicken β-actin
pOPING-ET vector Vector Map
Plasmid Resuspension Protocol:
1. Centrifuge at 5,000×g for 5 min.
2. Carefully open the tube and add 20 μl of sterile water to dissolve the DNA.
3. Close the tube and incubate for 10 minutes at room temperature.
4. Briefly vortex the tube and then do a quick spin to concentrate the liquid at the bottom. Speed is less than 5000×g.
5.Store the plasmid at -20 ℃.
pOPING-ET vector Sequence
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"
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