Basic Vector Information
- Vector Name:
- BE4-Gam
- Antibiotic Resistance:
- Ampicillin
- Length:
- 9444 bp
- Type:
- Mammalian Expression
- Replication origin:
- ori
- Copy Number:
- High Copy
- Promoter:
- CMV
- Cloning Method:
- Gibson Cloning
- 5' Primer:
- T7
- 3' Primer:
- M13R
BE4-Gam 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 ℃.
BE4-Gam vector Sequence
LOCUS BE4-Gam. 9444 bp DNA circular SYN 13-MAY-2021
DEFINITION Expresses BE4-Gam in mammalian cells.
ACCESSION .
VERSION .
KEYWORDS BE4-Gam.
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 9444)
AUTHORS Komor AC, Zhao KT, Packer MS, Gaudelli NM, Waterbury AL, Koblan LW,
Kim YB, Badran AH, Liu DR
TITLE Improved base excision repair inhibition and bacteriophage Mu Gam
protein yields C:G-to-T:A base editors with higher efficiency and
product purity.
JOURNAL Sci Adv. 2017 Aug 30;3(8):eaao4774. doi: 10.1126/sciadv.aao4774.
eCollection 2017 Aug.
PUBMED 28875174
REFERENCE 2 (bases 1 to 9444)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 9444)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Sci Adv.";
date: "2017-08-30"; pages: "
10.1126/sciadv.aao4774. eCollection 2017 Aug"
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..9444
/mol_type="other DNA"
/organism="synthetic DNA construct"
promoter 132..335
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
promoter 377..395
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 409..930
/gene="gam"
/label=gam
/note="Putative DNA ends protecting protein gam from
Escherichia phage Mu. Accession#: P06023"
CDS 979..1662
/label=APOBEC-1
/note="cytidine deaminase (C to U editing enzyme) from rat"
CDS 1759..5859
/label=Cas9(D10A)
/note="nickase mutant of the Cas9 endonuclease from the
Streptococcus pyogenes Type II CRISPR/Cas system"
CDS 5890..6138
/label=UGI
/note="uracil-DNA glycosylase inhibitor from a Bacillus
subtilis bacteriophage (Mol et al., 1995)"
CDS 6169..6417
/codon_start=1
/product="uracil-DNA glycosylase inhibitor from a Bacillus
subtilis bacteriophage (Mol et al., 1995)"
/label=UGI
/translation="TNLSDIIEKETGKQLVIQESILMLPEEVEEVIGNKPESDILVHTA
YDESTDENVMLLTSDAPEYKPWALVIQDSNGENKIKML"
CDS 6430..6450
/label=SV40 NLS
/note="nuclear localization signal of SV40 (simian virus
40) large T antigen"
CDS 6459..6476
/label=6xHis
/note="6xHis affinity tag"
polyA_signal 6505..6729
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
primer_bind complement(6800..6816)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind complement(6824..6840)
/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(6848..6878)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(6893..6914)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(7031..7048)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(7202..7790)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(7964..8821)
/label=AmpR
/note="beta-lactamase"
promoter complement(8822..8926)
/label=AmpR promoter
primer_bind complement(9005..9024)
/label=pRS-marker
/note="pRS vectors, use to sequence yeast selectable
marker"
enhancer 9196..9444
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
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