Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V011106 | pIRES2-EGFP | 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
- Vector Name:
- pIRES2-EGFP
- Antibiotic Resistance:
- Kanamycin
- Length:
- 5308 bp
- Type:
- Mammalian Expression Vectors
- Replication origin:
- ori
- Source/Author:
- Clontech
- Selection Marker:
- Neomycin/G418(Geneticin)
- Copy Number:
- High copy number
- Promoter:
- CMV
- Growth Strain(s):
- stbl3
- Growth Temperature:
- 37℃
pIRES2-EGFP 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
- Li BN, Li WD, Lin JT, Feng HG, Yuan ZQ. Construction and identification of pIRES2-LIF-NT-3 bicistronic eukaryotic expression vector. Genet Mol Res. 2014 Jun 18;13(2):4691-703. doi: 10.4238/2014.June.18.12. PMID: 25036519.
- Niu P, Huang CX, Zhao YQ, Yang B, Zhao QY, Wang T, Fan GH. [Recombinant plasmid pIRES2-EGFP-HCN2 improved pacing function in canine model of sick sinus syndrome]. Zhonghua Xin Xue Guan Bing Za Zhi. 2006 Dec;34(12):1126-30. Chinese. PMID: 17274909.
pIRES2-EGFP vector Sequence
LOCUS 40924_25681 5308 bp DNA circular SYN 01-JAN-1980
DEFINITION IRES-containing bicistronic vector for expressing a gene together
with EGFP.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 5308)
AUTHORS Clontech
TITLE Direct Submission
REFERENCE 2 (bases 1 to 5308)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
COMMENT The gene inserted into the MCS should contain start and stop codons.
FEATURES Location/Qualifiers
source 1..5308
/mol_type="other DNA"
/organism="synthetic DNA construct"
enhancer 61..364
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 365..568
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
misc_feature 600..665
/label=MCS
/note="multiple cloning site"
misc_feature 667..1253
/label=IRES2
/note="internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)"
CDS 1254..1970
/codon_start=1
/label=EGFP
/note="enhanced GFP"
/translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
EFVTAAGITLGMDELYK"
polyA_signal 2096..2217
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
rep_origin complement(2224..2679)
/direction=LEFT
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 2706..2810
/label=AmpR promoter
promoter 2812..3169
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
CDS 3204..3995
/codon_start=1
/label=NeoR/KanR
/note="aminoglycoside phosphotransferase"
/translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
polyA_signal 4230..4277
/label=HSV TK poly(A) signal
/note="herpes simplex virus thymidine kinase
polyadenylation signal (Cole and Stacy, 1985)"
rep_origin 4606..5194
/direction=RIGHT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"