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pVAX1-CMV-MCS-3×T2A-SV40-Hyg vector (V013492)

Price Information

Cat No. Plasmid Name Availability Add to cart
V013492 pVAX1-CMV-MCS-3×T2A-SV40-Hyg In stock, 1 week for quality controls

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

Basic Vector Information

Vector Name:
pVAX1-CMV-MCS-3×T2A-SV40-Hyg
Antibiotic Resistance:
Kanamycin
Length:
4771 bp
Type:
DNA vaccine, Protein expression
Replication origin:
ori
Host:
Mammalian cells
Selection Marker:
Hyg
Promoter:
CMV
Growth Strain(s):
DH5a
Growth Temperature:
37℃

pVAX1-CMV-MCS-3×T2A-SV40-Hyg vector Map

Copy Sequence View Map
pVAX1-CMV-MCS-3×T2A-SV40-Hyg4771 bp600120018002400300036004200CMV enhancerCMV promoterT7 promoterT2AbGH poly(A) signalSV40 promoterHygRSV40 poly(A) signalNeoR/KanRori

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.

pVAX1-CMV-MCS-3×T2A-SV40-Hyg vector Sequence

LOCUS       62056_22490        4771 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 4771)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..4771
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        36..415
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        416..619
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early
                     promoter"
     promoter        664..682
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             711..764
                     /codon_start=1
                     /label=T2A
                     /note="2A peptide from Thosea asigna virus capsid protein"
                     /translation="EGRGSLLTCGDVEENPGP"
     polyA_signal    946..1170
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     promoter        1280..1609
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             1658..2680
                     /codon_start=1
                     /label=HygR
                     /note="aminoglycoside phosphotransferase from E. coli"
                     /translation="MKKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRG
                     YVLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLP
                     ETELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQ
                     TVMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMF
                     GDSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDG
                     NFDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAK
                     E"
     polyA_signal    2813..2934
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     CDS             2998..3789
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     rep_origin      4118..4706
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"