Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V015023 | pCMV-Twinstrep-NK1R-mEGFP-FLAG-10×His-mCherry | 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:
- pCMV-Twinstrep-NK1R-mEGFP-FLAG-10×His-mCherry
- Antibiotic Resistance:
- Ampicillin
- Length:
- 9536 bp
- Type:
- Protein expression
- Replication origin:
- ori
- Host:
- Mammalian cells
- Promoter:
- CMV
- Growth Temperature:
- 37℃
pCMV-Twinstrep-NK1R-mEGFP-FLAG-10×His-mCherry 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-Twinstrep-NK1R-mEGFP-FLAG-10×His-mCherry vector Sequence
LOCUS 62056_7625 9536 bp DNA circular SYN 01-JAN-1980
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 9536)
AUTHORS .
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..9536
/mol_type="other DNA"
/organism="synthetic DNA construct"
rep_origin 102..556
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 582..686
/label=AmpR promoter
CDS 687..1544
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
rep_origin 1718..2306
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
mobile_element complement(2611..2835)
/label=Tn7R
/note="mini-Tn7 element (right end of the Tn7 transposon)"
CDS complement(2905..3435)
/codon_start=1
/label=GmR
/note="gentamycin acetyltransferase"
/translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPKFEQPRS
EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
EEVMHFDIDPSTAT"
promoter complement(3624..3652)
/label=Pc promoter
/note="class 1 integron promoter"
polyA_signal complement(4169..4217)
/label=HSV TK poly(A) signal
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
CDS complement(4291..4998)
/codon_start=1
/label=mCherry
/note="monomeric derivative of DsRed fluorescent protein
(Shaner et al., 2004)"
/translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
EGRHSTGGMDELYK"
promoter complement(5031..5140)
/label=p10 promoter
/note="baculovirus promoter for expression in insect cells"
promoter 5185..5276
/label=polyhedrin promoter
/note="promoter for the baculovirus polyhedrin gene"
enhancer 5457..5835
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 5836..6039
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
intron 6175..6307
/label=chimeric intron
/note="chimera between introns from human beta-globin and
immunoglobulin heavy chain genes"
CDS 6419..6442
/codon_start=1
/label=Strep-Tag II
/note="peptide that binds Strep-Tactin(R), an engineered
form
of streptavidin"
/translation="WSHPQFEK"
CDS 6479..6502
/codon_start=1
/label=Strep-Tag II
/note="peptide that binds Strep-Tactin(R), an engineered
form
of streptavidin"
/translation="WSHPQFEK"
CDS 7565..7585
/codon_start=1
/label=TEV site
/note="tobacco etch virus (TEV) protease recognition and
cleavage site"
/translation="ENLYFQG"
CDS 7604..8320
/codon_start=1
/label=mEGFP
/note="enhanced GFP with monomerizing A206K mutation
(Zacharias et al., 2002)"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
CDS 8321..8344
/codon_start=1
/label=FLAG
/note="FLAG(R) epitope tag, followed by an enterokinase
cleavage site"
/translation="DYKDDDDK"
CDS 8345..8371
/codon_start=1
/label=9xHis
/note="9xHis affinity tag"
/translation="HHHHHHHHH"
misc_feature 8405..8993
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
polyA_signal 9126..9260
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
mobile_element complement(9289..9454)
/label=Tn7L
/note="mini-Tn7 element (left end of the Tn7 transposon)"