pcDNA3-EGFP vector (V018125)

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Price Information

Cat No.	Plasmid Name	Availability	Buy one, get one free! (?)
V018125	pcDNA3-EGFP	In stock, 1 week for quality controls

Two tubes of lyophilized plasmid will be delivered, each tube is about 5µg.

Basic Vector Information

The pcDNA3-EGFP is a mammalian expression plasmid containing the enhanced green fluorescent protein (EGFP) gene driven by the CMV promoter for high-level expression in mammalian cells. It carries ampicillin resistance for bacterial selection and neomycin/G418 resistance for stable selection in eukaryotic cells. This plasmid is widely used as a reporter construct and for creating EGFP fusion proteins.

Vector Name:: pcDNA3-EGFP

Antibiotic Resistance:: Ampicillin

Length:: 6159 bp

Type:: Mammalian Expression Vectors

Source/Author:: Doug Golenbock

Copy Number:: High copy number

Promoter:: CMV

Growth Temperature:: 37℃

pcDNA3-EGFP vector Map

Copy Sequence View 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

Martins CS, Costa DVS, Lima BB, Leitäo RFC, Freire GE, Silva GFM, Pacífico DM, Abreu JG, Brito GAC. Clostridioides difficile Toxin A-Induced Wnt/β-Catenin Pathway Inhibition Is Mediated by Rac1 Glucosylation. Front Microbiol. 2020 Aug 28;11:1998. doi: 10.3389/fmicb.2020.01998. PMID: 32983019; PMCID: PMC7483921.

pcDNA3-EGFP vector Sequence

LOCUS       Exported                6159 bp DNA     circular SYN 14-APR-2025
DEFINITION  Mammalian expression vector for cloning your gene fused to EGFP.
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3-EGFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6159)
  TITLE     Fluorescent Protein Plasmids
REFERENCE   2  (bases 1 to 6159)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6159
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..380
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        381..584
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     535..555
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     629..648
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        629..647
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             743..1462
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             743..1459
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(788..809)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1049..1068)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1396..1417
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     promoter        complement(1478..1496)
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     complement(1479..1496)
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     primer_bind     complement(1516..1533)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1522..1746
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      1792..2220
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1879..1898)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2089..2110
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2229..2249)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        2234..2563
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      2414..2549
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2476..2495
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             2630..3424
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(2684..2703)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     3294..3313
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    3598..3719
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3635..3654)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3689..3708
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3768..3784)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3768..3784)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3781..3803)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3792..3808
                     /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(3816..3846)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3861..3882
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3999..4016)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4170..4758)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4250..4269)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4929..5789)
                     /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"
     primer_bind     5552..5571
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5790..5894)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5969..5988)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"