Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V001465 | pCXWB-EBNA1 | 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
Non-replicating episomal expression of EBNA1, which enhances transfection efficiency and expression from episomal plasmids
- Vector Name:
- pCXWB-EBNA1
- Antibiotic Resistance:
- Ampicillin
- Length:
- 7037 bp
- Type:
- Non-replicating episomal expression of EBNA1
- Replication origin:
- ori
- Source/Author:
- Shinya Yamanaka
- Copy Number:
- High copy number
- Promoter:
- CAG
- Cloning Method:
- Enzyme Cut
- Growth Strain(s):
- Stbl3
- Growth Temperature:
- 37℃
- Expression Method:
- Transient
pCXWB-EBNA1 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
- Xing K, Cui Y, Luan J, Zhou X, Shi L, Han J. Establishment of a human trisomy 18 induced pluripotent stem cell line from amniotic fluid cells. Intractable Rare Dis Res. 2018 May;7(2):94-99.
pCXWB-EBNA1 vector Sequence
LOCUS 40924_14060 7037 bp DNA circular SYN 07-AUG-2018
DEFINITION Non-replicating episomal expression of EBNA1, which enhances
transfection efficiency and expression from episomal plasmids.
ACCESSION .
VERSION .
KEYWORDS .
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 7037)
AUTHORS Okita K, Yamakawa T, Matsumura Y, Sato Y, Amano N, Watanabe A,
Goshima N, Yamanaka S
TITLE An Efficient Non-viral Method to Generate Integration-Free Human iPS
Cells from Cord Blood and Peripheral Blood Cells.
JOURNAL Stem Cells. 2012 Nov 29. doi: 10.1002/stem.1293.
PUBMED 23193063
REFERENCE 2 (bases 1 to 7037)
TITLE Direct Submission
REFERENCE 3 (bases 1 to 7037)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Stem Cells.
2012 Nov 29. doi: 10.1002/stem.1293."
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..7037
/mol_type="other DNA"
/organism="synthetic DNA construct"
enhancer 3..382
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 384..660
/label=chicken beta-actin promoter
intron 661..1677
/label=chimeric intron
/note="chimera between introns from chicken beta-actin and
rabbit beta-globin"
primer_bind 1685..1704
/label=pCAG-F
/note="Rabbit beta-globin intron, for pCAG plasmids,
forward primer"
CDS 1747..3669
/codon_start=1
/label=EBNA1
/note="Epstein-Barr nuclear antigen 1, also known as
EBNA-1"
/translation="MSDEGPGTGPGNGLGEKGDTSGPEGSGGSGPQRRGGDNHGRGRGR
GRGRGGGRPGAPGGSGSGPRHRDGVRRPQKRPSCIGCKGTHGGTGAGAGAGGAGAGGAG
AGGGAGAGGGAGGAGGAGGAGAGGGAGAGGGAGGAGGAGAGGGAGAGGGAGGAGAGGGA
GGAGGAGAGGGAGAGGGAGGAGAGGGAGGAGGAGAGGGAGAGGAGGAGGAGAGGAGAGG
GAGGAGGAGAGGAGAGGAGAGGAGAGGAGGAGAGGAGGAGAGGAGGAGAGGGAGGAGAG
GGAGGAGAGGAGGAGAGGAGGAGAGGAGGAGAGGGAGAGGAGAGGGGRGRGGSGGRGRG
GSGGRGRGGSGGRRGRGRERARGGSRERARGRGRGRGEKRPRSPSSQSSSSGSPPRRPP
PGRRPFFHPVGEADYFEYHQEGGPDGEPDVPPGAIEQGPADDPGEGPSTGPRGQGDGGR
RKKGGWFGKHRGQGGSNPKFENIAEGLRALLARSHVERTTDEGTWVAGVFVYGGSKTSL
YNLRRGTALAIPQCRLTPLSRLPFGMAPGPGPQPGPLRESIVCYFMVFLQTHIFAEVLK
DAIKDLVMTKPAPTCNIRVTVCSFDDGVDLPPWFPPMVEGAAAEGDDGDDGDEGGDGDE
GEEGQE"
misc_feature 3716..4304
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
primer_bind complement(4394..4413)
/label=Bglob-pA-R
/note="Rabbit beta-globin polyA region, reverse primer"
polyA_signal 4459..4514
/label=beta-globin poly(A) signal
/note="rabbit beta-globin polyadenylation signal (Gil and
Proudfoot, 1987)"
primer_bind complement(4513..4532)
/label=rbglobpA-R
/note="Rabbit beta-globin polyA, reverse primer. Also
called rb-glob-pA-term-R"
primer_bind complement(4875..4891)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
protein_bind complement(4899..4915)
/label=lac operator
/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(4923..4953)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(4968..4989)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
primer_bind complement(5114..5131)
/label=L4440
/note="L4440 vector, forward primer"
rep_origin complement(5285..5873)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(6047..6904)
/codon_start=1
/label=AmpR
/note="beta-lactamase"
/translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
LIKHW"
promoter complement(6905..7009)
/label=AmpR promoter
promoter 7037
/label=CAG
/note="CMV early enhancer fused to modified chicken
beta-actin promoter"