Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V001615 | pBudCE4.1 | 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
The pBudCE4.1 vector is designed for the independent expression of two genes from a single plasmid in mammalian cells. Using pBudCE4.1 to generate stable mammalian expression cell lines ensures that there is an equivalent copy number of each gene in the cell. This can eliminate variable expression due to differences in gene copy number. pBudCE4.1 provides expression cassettes withthe following features:
- Vector Name:
- pBudCE4.1
- Antibiotic Resistance:
- Zeocin
- Length:
- 4595 bp
- Type:
- Mammalian Expression Vectors (Dual Promoter)
- Replication origin:
- ori
- Selection Marker:
- Zeocin
- Promoter:
- EF-1α
- Cloning Method:
- Enzyme digestion and ligation
- 5' Primer:
- CMVPro-F: 5'd[TAATACGACTCACTATAGGG]3' EF1aPro-F:5'd[TCAAGCCTCAGACAGTGGTTC]3'
- Fusion Tag:
- 6xHis, c-Myc, V5
- Growth Strain(s):
- stbl3
- Growth Temperature:
- 37℃
pBudCE4.1 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
- Zhao RL, Zhang XM, Jia LN, Song W, Sun YL, Meng XY, Peng XX. pNNS-Conjugated Chitosan Mediated IGF-1 and miR-140 Overexpression in Articular Chondrocytes Improves Cartilage Repair. Biomed Res Int. 2019 Mar 21;2019:2761241.
pBudCE4.1 vector Sequence
LOCUS 40924_7606 4595 bp DNA circular SYN 13-JAN-2022
DEFINITION synthetic circular DNA.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 4595)
TITLE Direct Submission
REFERENCE 2 (bases 1 to 4595)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..4595
/mol_type="other DNA"
/organism="synthetic DNA construct"
enhancer 10..389
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 390..593
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
promoter 638..656
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
CDS 719..748
/codon_start=1
/label=Myc
/note="Myc (human c-Myc proto-oncogene) epitope tag"
/translation="EQKLISEEDL"
CDS 764..781
/codon_start=1
/label=6xHis
/note="6xHis affinity tag"
/translation="HHHHHH"
polyA_signal complement(800..933)
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
CDS complement(1066..1437)
/codon_start=1
/label=BleoR
/note="antibiotic-binding protein"
/translation="MAKLTSAVPVLTARDVAGAVEFWTDRLGFSRDFVEDDFAGVVRDD
VTLFISAVQDQVVPDNTLAWVWVRGLDELYAEWSEVVSTNFRDASGPAMTEIGEQPWGR
EFALRDPAGNCVHFVAEEQD"
promoter complement(1456..1503)
/label=EM7 promoter
/note="synthetic bacterial promoter"
promoter complement(1551..1867)
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
promoter 1889..3051
/label=EF-1-alpha promoter
/note="strong constitutive promoter for human elongation
factor EF-1-alpha"
CDS 3127..3168
/codon_start=1
/label=V5 tag
/note="epitope tag from simian virus 5"
/translation="GKPIPNPLLGLDST"
CDS 3178..3195
/codon_start=1
/label=6xHis
/note="6xHis affinity tag"
/translation="HHHHHH"
polyA_signal 3224..3448
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin complement(3580..4168)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"