Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V001682 | pCWori | In stock, instant shipping |
Buy one, get one free! |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
For e.coli protein expression.
- Vector Name:
- pCWori
- Antibiotic Resistance:
- Ampicillin
- Length:
- 5438 bp
- Type:
- Prokaryotic Expression Vectors
- Replication origin:
- ori
- Promoter:
- tac/lac
pCWori 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.
References
- Muchmore DC, McIntosh LP, Russell CB, Anderson DE, Dahlquist FW. Expression and nitrogen-15 labeling of proteins for proton and nitrogen-15 nuclear magnetic resonance. Methods Enzymol. 1989;177:44-73.
- Fisher CW, Caudle DL, Martin-Wixtrom C, Quattrochi LC, Tukey RH, Waterman MR, Estabrook RW. High-level expression of functional human cytochrome P450 1A2 in Escherichia coli. FASEB J. 1992 Jan 6;6(2):759-64.
- Gerber NC, Ortiz de Montellano PR. Neuronal nitric oxide synthase. Expression in Escherichia coli, irreversible inhibition by phenyldiazene, and active site topology. J Biol Chem. 1995 Jul 28;270(30):17791-6. doi: 10.1074/jbc.270.30.17791. PMID: 7543092.
- Kawashima H, Kusunose E, Thompson CM, Strobel HW. Protein expression, characterization, and regulation of CYP4F4 and CYP4F5 cloned from rat brain. Arch Biochem Biophys. 1997 Nov 1;347(1):148-54. doi: 10.1006/abbi.1997.0342. PMID: 9344476.
pCWori vector Sequence
LOCUS Exported 5438 bp DNA circular SYN 31-AUG-2024
DEFINITION Exported.
ACCESSION V001682
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5438)
TITLE Direct Submission
REFERENCE 2 (bases 1 to 5438)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 5438)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..5438
/mol_type="other DNA"
/organism="synthetic DNA construct"
source 644..649
/mol_type="other DNA"
/organism="synthetic DNA construct"
source 1126..1131
/mol_type="other DNA"
/organism="synthetic DNA construct"
CDS 21..407
/gene="cheY"
/label=Chemotaxis protein CheY
/note="Chemotaxis protein CheY from Escherichia coli
(strain K12). Accession#: P0AE67"
primer_bind complement(465..481)
/label=SK primer
/note="common sequencing primer, one of multiple similar
variants"
rep_origin 668..1123
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 1156..1260
/label=AmpR promoter
CDS 1261..2118
/label=AmpR
/note="beta-lactamase"
rep_origin 2292..2880
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
misc_feature complement(3066..3207)
/label=bom
/note="basis of mobility region from pBR322"
CDS complement(3616..3789)
/label=lacZ-alpha
/note="LacZ-alpha fragment of beta-galactosidase"
protein_bind complement(3809..3825)
/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(3833..3863)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(3878..3899)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
CDS complement(3915..4994)
/label=lacI
/note="lac repressor"
promoter complement(4995..5072)
/label=lacIq promoter
/note="In the lacIq allele, a single base change in the
promoter boosts expression of the lacI gene about 10-fold."
protein_bind 5124..5145
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 5160..5190
/label=lac UV5 promoter
/note="E. coli lac promoter with an 'up' mutation"
protein_bind 5198..5214
/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 5269..5297
/label=tac promoter
/note="strong E. coli promoter; hybrid between the trp and
lac UV5 promoters"
protein_bind 5305..5321
/label=lac repressor encoded by lacI binding site
/bound_moiety="lac repressor encoded by lacI"
/note="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)."
misc_feature 5364..5392
/label=tac promoter
/note="tac promoter"
/note="strong E. coli promoter; hybrid between the trp and
lac UV5 promoters"
protein_bind 5400..5416
/label=lac repressor encoded by lacI binding site
/bound_moiety="lac repressor encoded by lacI"
/note="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)."