Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V011361 | p3xFLAG-CMV-14 | 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
p3xFLAG-CMV-14 is a mammalian expression vector designed for the constitutive expression of a target protein in mammalian cells.
p3xFLAG-CMV-14 features the strong cytomegalovirus (CMV) promoter to drive high levels of gene expression. The multiple cloning site (MCS) contains three FLAG tags, allowing for easy detection and immunoprecipitation of the expressed protein. Additionally, the vector includes the SV40 origin of replication, enabling efficient replication in mammalian cells. The ampicillin resistance gene confers resistance to ampicillin, facilitating selection of transformed cells.
p3xFLAG-CMV-14 is commonly used in protein overexpression studies, subcellular localization analysis, and protein-protein interaction assays in mammalian cell culture systems.
- Vector Name:
- p3xFLAG-CMV-14
- Antibiotic Resistance:
- Ampicillin
- Length:
- 6310 bp
- Type:
- Mammalian Expression Vectors
- Replication origin:
- ori
- Source/Author:
- Sigma-Aldrich
- Selection Marker:
- Neomycin/G418(Geneticin)
- Copy Number:
- High copy number
- Promoter:
- CMV
- Cloning Method:
- Enzyme digestion and ligation
- 5' Primer:
- M13 fwd
- 3' Primer:
- M13 rev
- Growth Strain(s):
- DH10B
- Growth Temperature:
- 37℃
p3xFLAG-CMV-14 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.
p3xFLAG-CMV-14 vector Sequence
LOCUS p3xFLAG-CMV-14. 6310 bp DNA circular SYN 01-JAN-1980
DEFINITION Vector with a neomycin resistance marker for expression of
C-terminally 3xFLAG-tagged proteins in mammalian cells.
ACCESSION .
VERSION .
KEYWORDS p3xFLAG-CMV-14
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 6310)
AUTHORS Sigma-Aldrich
TITLE Direct Submission
REFERENCE 2 (bases 1 to 6310)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..6310
/mol_type="other DNA"
/organism="synthetic DNA construct"
primer_bind 141..157
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
enhancer 318..697
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 698..901
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
misc_feature 924..990
/label=MCS
/note="MCS"
/note="multiple cloning site"
CDS 993..1058
/label=3xFLAG
/note="three tandem FLAG(R) epitope tags, followed by an
enterokinase cleavage site"
polyA_signal 1069..1691
/label=hGH poly(A) signal
/note="human growth hormone polyadenylation signal"
promoter 1733..2030
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
CDS 2084..2875
/label=NeoR/KanR
/note="aminoglycoside phosphotransferase"
polyA_signal 3530..3664
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
promoter complement(3670..3688)
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
primer_bind complement(3702..3718)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind complement(3726..3742)
/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(3750..3780)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(3795..3816)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(4104..4692)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(4866..5723)
/label=AmpR
/note="beta-lactamase"
promoter complement(5724..5828)
/label=AmpR promoter
rep_origin 5855..6310
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"