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pVitro2-neo-mcs vector (V012767)

Price Information

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V012767 pVitro2-neo-mcs In stock, instant shipping

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

Basic Vector Information

Expression plasmids pVITRO2-MCS are developed mainly for in vitro studies. pVITRO2-MCS allows the ubiquitous and constitutive co-expression of two genes of interest. pVITRO2-MCS plasmids carry two human ferritin composite promoters, FerH (heavy chain) and FerL (light chain). To eliminate the iron regulation, their 5’UTRs have been replaced by the 5’UTR of the mouse and chimpanzee EF-1α genes. pVITRO2-MCS plasmids are available with different selectable markers that are active both in E. coli and mammalian cells. pVITRO2-MCS plasmids contain two multiple cloning sites (MCS) for the convenient cloning of your cDNAs of interest.

Vector Name:
pVitro2-neo-mcs
Antibiotic Resistance:
Kanamycin
Length:
6125 bp
Type:
Mammalian Expression Vectors
Replication origin:
ori
Source/Author:
Invivogen
Selection Marker:
Neomycin/G418(Geneticin)
Copy Number:
High copy number
Promoter:
CMV
Cloning Method:
Enzyme Cut
5' Primer:
EF-1a-F：TCAAGCCTCAGACAGTGGTTC
Expression Method:
Constiutive, Stable / Transient

pVitro2-neo-mcs vector Map

Copy Sequence View Map
pVitro2-neo-mcs6125 bp30060090012001500180021002400270030003300360039004200450048005100540057006000CMV enhancerEF-1-alpha intron ASV40 poly(A) signaloriSV40 enhancermEF-1-alpha intronFMDV IRESEM7 promoterNeoR/KanREF-1-alpha poly(A) signal

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.

pVitro2-neo-mcs vector Sequence

LOCUS       40924_45993        6125 bp DNA     circular SYN 18-SEP-2021
DEFINITION  synthetic circular DNA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6125)
  TITLE     Direct Submission
REFERENCE   2  (bases 1 to 6125)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..6125
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        9..312
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          700..1644
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     polyA_signal    complement(1728..1849)
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     rep_origin      2037..2625
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     enhancer        2747..2938
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     intron          3145..4092
                     /label=mEF-1-alpha intron
                     /note="intron upstream of the start codon of mouse 
                     EF-1-alpha"
     misc_feature    4167..4611
                     /label=FMDV IRES
                     /note="internal ribosome entry site (IRES) of the
                     foot-and-mouth disease virus"
     promoter        4644..4691
                     /label=EM7 promoter
                     /note="synthetic bacterial promoter"
     CDS             4711..5502
                     /codon_start=1
                     /label=NeoR/KanR
                     /note="aminoglycoside phosphotransferase"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     polyA_signal    5552..6124
                     /label=EF-1-alpha poly(A) signal
                     /note="human EF-1-alpha polyadenylation signal"