Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V008563 | pCEV-G4-Km | In stock, 1 week for quality controls |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
- Vector Name:
- pCEV-G4-Km
- Antibiotic Resistance:
- Ampicillin
- Length:
- 6574 bp
- Type:
- Cloning and expression vector
- Replication origin:
- ori
- Host:
- Yeast
- Source/Author:
- Vickers CE, Bydder SF, Zhou Y, Nielsen LK.
- Promoter:
- TEF1
pCEV-G4-Km vector Vector Map
Plasmid 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 ℃.
6. The concentration of plasmid re-measurement sometimes differs from the nominal value, which may be due to the position of the lyophilized plasmid in the tube, the efficiency of the re-dissolution, the measurement bias, and adsorption on the wall of the tube, therefore, it is recommended to transform and extract the plasmid before using it
General Plasmid Transform Protocol
1. Take one 100μl of the competent cells and thaw it on ice for 10min, add 2μl of plasmid, then ice bath for 30min, then heat-shock it at 42℃ for 60s, do not stir, and then ice bath for 2min.
2. Add 900μl of LB liquid medium without antibiotics, and incubate at 37℃ for 45min (30℃ for 1-1.5 hours) with 180rpm shaking.
3. Centrifuge at 6000rpm for 5min, leave only 100μl of supernatant to resuspend the bacterial precipitate and spread it onto the target plasmid-resistant LB plate.
4. Invert the plate and incubate at 37℃ for 14h, or at 30℃ for 20h.
5. Pick a single colony into LB liquid medium, add the corresponding antibiotics, incubate at 220rpm for 14h, and extract the plasmid according to the experimental needs and the instructions of the plasmid extraction kit.
pCEV-G4-Km vector Sequence
LOCUS 40924_10436 6574 bp DNA circular SYN 17-DEC-2018
DEFINITION Cloning and expression vector pCEV-G4-Km, complete sequence.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6574)
AUTHORS Vickers CE, Bydder SF, Zhou Y, Nielsen LK.
TITLE Dual gene expression cassette vectors with antibiotic selection
markers for engineering in Saccharomyces cerevisiae
JOURNAL Microb. Cell Fact. 12 (1), 96 (2013)
PUBMED 24161108
REFERENCE 2 (bases 1 to 6574)
AUTHORS Bydder SF, Vickers CE.
TITLE Direct Submission
JOURNAL Submitted (08-JUL-2013) AIBN, The University of Queensland, Crn
College and Cooper Roads (Bldg 75), St Lucia, QLD 4072, Australia
REFERENCE 3 (bases 1 to 6574)
TITLE Direct Submission
REFERENCE 4 (bases 1 to 6574)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Microb.
Cell Fact."; date: "2013"; volume: "12"; issue: "1"; pages: "96"
COMMENT SGRef: number: 2; type: "Journal Article"; journalName: "Submitted
(08-JUL-2013) AIBN, The University of Queensland, Crn College and
Cooper Roads (Bldg 75), St Lucia, QLD 4072, Australia"
COMMENT SGRef: number: 3; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..6574
/mol_type="other DNA"
/organism="synthetic DNA construct"
terminator complement(2..167)
/label=ADH1 terminator
/note="transcription terminator for the S. cerevisiae
alcohol dehydrogenase 1 (ADH1) gene"
CDS complement(315..338)
/label=FLAG
/note="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
regulatory complement(352..937)
/label=HXT7 promoter from Saccharomyces cerevisiae
/note="HXT7 promoter from Saccharomyces cerevisiae"
/regulatory_class="promoter"
promoter 946..1353
/label=TEF1 promoter
/note="promoter for EF-1-alpha"
promoter 1373..1391
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 1409..1438
/label=Myc
/note="Myc (human c-Myc proto-oncogene) epitope tag"
terminator 1468..1657
/label=CYC1 terminator
/note="transcription terminator for CYC1"
rep_origin complement(1900..2488)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(2662..3519)
/label=AmpR
/note="beta-lactamase"
promoter complement(3520..3624)
/label=AmpR promoter
rep_origin complement(3651..4993)
/direction=LEFT
/label=2u ori
/note="yeast 2u plasmid origin of replication"
protein_bind 5013..5046
/label=loxP
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (ATGTATGC) (Shaw et al., 2021)."
gene 5101..6457
/label=kanMX
/note="yeast selectable marker conferring kanamycin
resistance (Wach et al., 1994)"
misc_recomb 6520..6553
/label=loxP recognition sequence for Cre recombinase
/note="loxP recognition sequence for Cre recombinase"
protein_bind complement(6520..6553)
/label=loxP
/bound_moiety="Cre recombinase"
/note="Cre-mediated recombination occurs in the 8-bp core
sequence (GCATACAT)."