Price Information
| Cat No. | Plasmid Name | Availability | Buy one, get one free! (?) |
|---|---|---|---|
| V018054 | pCMV-TurboID-3×FLAG-Neo | In stock, instant shipping |
Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.
Basic Vector Information
The pCMV - TurboID - 3×FLAG - Neo plasmid is for studying protein - protein interactions. It uses the pCMV promoter to express the TurboID - target protein fusion in mammalian cells. TurboID labels interacting proteins, the 3×FLAG tag aids fusion protein detection and purification, and the Neo gene screens stable transfected cell lines, offering a powerful tool for protein function and signaling pathway research.
- Vector Name:
- pCMV-TurboID-3×FLAG-Neo
- Antibiotic Resistance:
- Ampicillin
- Length:
- 6433 bp
- Type:
- Protein expression
- Replication origin:
- ori
- Host:
- Mammalian cells
- Selection Marker:
- Neo/G418
- Promoter:
- CMV
- 5' Primer:
- CMV-F
- Growth Strain(s):
- DH10B
- Growth Temperature:
- 37℃
pCMV-TurboID-3×FLAG-Neo 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.
pCMV-TurboID-3×FLAG-Neo vector Sequence
LOCUS . 6433 bp DNA circular UNK 01-JAN-1980
DEFINITION synthetic circular DNA.
ACCESSION
VERSION
KEYWORDS .
SOURCE .
ORGANISM .
.
FEATURES Location/Qualifiers
enhancer 235..614
/label="CMV enhancer"
/note="human cytomegalovirus immediate early enhancer"
promoter 615..818
/label="CMV promoter"
/note="human cytomegalovirus (CMV) immediate early
promoter"
promoter 863..881
/label="T7 promoter"
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 964..1923
/label="TurboID"
/note="mutant of the E. coli biotin ligase BioID engineered
for improved catalytic efficiency (Branon et al., 2018)"
CDS 1930..1995
/label="3xFLAG"
/note="three tandem FLAG® epitope tags, followed by an
enterokinase cleavage site"
polyA_signal 2033..2257
/label="bGH poly(A) signal"
/note="bovine growth hormone polyadenylation signal"
rep_origin 2303..2731
/label="f1 ori"
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 2745..3074
/label="SV40 promoter"
/note="SV40 enhancer and early promoter"
CDS 3141..3932
/label="NeoR/KanR"
/note="aminoglycoside phosphotransferase"
polyA_signal 4109..4242
/label="SV40 poly(A) signal"
/note="SV40 polyadenylation signal"
primer_bind complement(4279..4295)
/label="M13 rev"
/note="common sequencing primer, one of multiple similar
variants"
protein_bind complement(4303..4319)
/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-β-D-thiogalactopyranoside (IPTG)."
promoter complement(4327..4357)
/label="lac promoter"
/note="promoter for the E. coli lac operon"
protein_bind complement(4372..4393)
/label="CAP binding site"
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(4681..5266)
/label="ori"
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(5440..6297)
/label="AmpR"
/note="β-lactamase"
promoter complement(6298..6402)
/label="AmpR promoter"