Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V001285 | pDRGFP | 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
In vivo homologous recombination substrate composed of two differentially mutated GFP genes oriented as direct repeats and separated by a drug selection marker.
- Vector Name:
- pDRGFP
- Antibiotic Resistance:
- Ampicillin
- Length:
- 8645 bp
- Type:
- Mammalian Expression
- Selection Marker:
- Puromycin
- Copy Number:
- High Copy
- Cloning Method:
- Restriction Enzyme
- 5' Primer:
- None
- Growth Strain(s):
- DH10B
- Growth Temperature:
- 37℃
pDRGFP 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.
pDRGFP vector Sequence
LOCUS Exported 8645 bp DNA circular SYN 30-OCT-2024
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 8645)
AUTHORS .
TITLE Direct Submission
FEATURES Location/Qualifiers
source 1..8645
/mol_type="other DNA"
/organism="synthetic DNA construct"
enhancer 4..383
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 385..660
/label=chicken beta-actin promoter
intron 661..1669
/label=chimeric intron
/note="chimera between introns from chicken beta-actin and
rabbit beta-globin"
CDS 1754..1774
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 1796..1816
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 1829..2017
/codon_start=1
/label=zinc finger domain
/translation="GNTSGVLSTPKAKRAKHPPGTEKPRSRSQSEQPATCPICYAVIRQ
SRNLRRHLELRHFAKPGV"
CDS 2018..2755
/codon_start=1
/label=SceGFP
/translation="DPPVATMVSKGEELFTGVVPILVELDGDVNGHKFSVSG*G*QGNT
YGKLTLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERT
IFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADK
QKNGIKVNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKR
DHMVLLEFVTAAGITLGMDELYK"
polyA_signal 2878..2999
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
CDS complement(3660..4259)
/codon_start=1
/gene="pac from Streptomyces alboniger"
/product="puromycin N-acetyltransferase"
/label=PuroR
/note="confers resistance to puromycin"
/translation="MTEYKPTVRLATRDDVPRAVRTLAAAFADYPATRHTVDPDRHIER
VTELQELFLTRVGLDIGKVWVADDGAAVAVWTTPESVEAGAVFAEIGPRMAELSGSRLA
AQQQMEGLLAPHRPKEPAWFLATVGVSPDHQGKGLGSAVVLPGVEAAERAGVPAFLETS
APRNLPFYERLGFTVTADVEVPEGPRTWCMTRKPGA"
promoter complement(4278..4775)
/label=PGK promoter
/note="mouse phosphoglycerate kinase 1 promoter"
polyA_signal 4928..4983
/label=beta-globin poly(A) signal
/note="rabbit beta-globin polyadenylation signal (Gil and
Proudfoot, 1987)"
CDS 5345..5365
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5387..5407
/codon_start=1
/product="nuclear localization signal of SV40 (simian virus
40) large T antigen"
/label=SV40 NLS
/translation="PKKKRKV"
CDS 5420..5608
/codon_start=1
/label=zinc finger domain
/translation="GNTSGVLSTPKAKRAKHPPGTEKPRSRSQSEQPATCPICYAVIRQ
SRNLRRHLELRHFAKPGV"
CDS 5609..6142
/codon_start=1
/label=iGFP
/translation="DPPVATMVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDAT
YGKLTLKFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERT
IFFKDDGNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADK
QKNGIKVNFNKSLA"
regulatory 5621..5630
/label=Kozak sequence
/note="vertebrate consensus sequence for strong initiation
of translation (Kozak, 1987)"
/regulatory_class="other"
primer_bind complement(6148..6164)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind 6172..6188
/label=lac operator
/bound_moiety="lac repressor encoded by lacI"
/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(6196..6226)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind 6241..6262
/label=CAP binding site
/bound_moiety="E. coli catabolite activator protein"
/note="CAP binding activates transcription in the presence
of cAMP."
promoter 6321..6516
/label=SV40 promoter
/note="SV40 early promoter"
rep_origin 6367..6502
/label=SV40 ori
/note="SV40 origin of replication"
polyA_signal 6522..6656
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
rep_origin complement(6895..7483)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(7654..8514)
/codon_start=1
/gene="bla"
/product="beta-lactamase"
/label=AmpR
/note="confers resistance to ampicillin, carbenicillin, and
related antibiotics"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(8515..8619)
/gene="bla"
/label=AmpR promoter