Price Information
| Cat No. | Plasmid Name | Availability | Add to cart |
|---|---|---|---|
| V017494 | pCBC-DT3T4 | 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
Bacterial Resistance(s)Chloramphenicol, 25 μg/mL. Used as CRISPR/Cas based plant genome editing and gene regulation; used as template for making expression cassette with multiple gRNA target sites.
- Vector Name:
- pCBC-DT3T4
- Antibiotic Resistance:
- Chloramphenicol
- Length:
- 3656 bp
- Type:
- Plant Expression, CRISPR
- Source/Author:
- Qi-Jun Chen
- Copy Number:
- High copy number
- Promoter:
- U6
- Growth Strain(s):
- DH5alpha
- Growth Temperature:
- 37℃
pCBC-DT3T4 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.
pCBC-DT3T4 vector Sequence
LOCUS Exported 3656 bp DNA circular SYN 29-JUN-2024
DEFINITION CRISPR/Cas based plant genome editing and gene regulation; used as
template for making expression cassette with multiple gRNA target
sites.
ACCESSION .
VERSION .
KEYWORDS pCBC-DT3T4
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 3656)
AUTHORS Xing HL, Dong L, Wang ZP, Zhang HY, Han CY, Liu B, Wang XC, Chen QJ
TITLE A CRISPR/Cas9 toolkit for multiplex genome editing in plants.
JOURNAL BMC Plant Biol. 2014 Nov 29;14(1):327.
PUBMED 25432517
REFERENCE 2 (bases 1 to 3656)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "BMC Plant
Biol."; date: "2014-11-29"; volume: "14(1)"; pages: "327"
FEATURES Location/Qualifiers
source 1..3656
/mol_type="other DNA"
/organism="synthetic DNA construct"
primer_bind 235..254
/label=CAT-R
/note="Chloramphenicol resistance gene, reverse primer"
CDS complement(669..971)
/codon_start=1
/gene="ccdB"
/product="CcdB, a bacterial toxin that poisons DNA gyrase"
/label=ccdB
/note="Plasmids containing the ccdB gene cannot be
propagated in standard E. coli strains."
/translation="QFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDKV
SRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
primer_bind complement(771..790)
/label=ccdB-fwd
/note="ccdB gene, forward primer"
primer_bind 1094..1116
/label=M13/pUC Forward
/note="In lacZ gene"
primer_bind 1108..1125
/label=M13 Forward
/note="In lacZ gene. Also called M13-F20 or M13 (-21)
Forward"
primer_bind 1109..1125
/label=M13 fwd
/note="common sequencing primer, one of multiple similar
variants"
primer_bind 1132..1151
/label=T7
/note="T7 promoter, forward primer"
promoter 1132..1150
/label=T7 promoter
/note="promoter for bacteriophage T7 RNA polymerase"
misc_RNA complement(1808..1883)
/label=gRNA scaffold
/note="guide RNA scaffold for the Streptococcus pyogenes
CRISPR/Cas9 system"
promoter complement(1954..1972)
/label=SP6 promoter
/note="promoter for bacteriophage SP6 RNA polymerase"
primer_bind complement(1955..1972)
/label=SP6
/note="SP6 promoter, forward primer"
primer_bind complement(1990..2006)
/label=M13 rev
/note="common sequencing primer, one of multiple similar
variants"
primer_bind complement(1990..2006)
/label=M13 Reverse
/note="In lacZ gene. Also called M13-rev"
primer_bind complement(2003..2025)
/label=M13/pUC Reverse
/note="In lacZ gene"
promoter complement(2045..2092)
/label=EM7 promoter
/note="synthetic bacterial promoter "
primer_bind complement(2103..2126)
/label=pENTR-F
/note="pENTR vectors, forward primer"
terminator 2134..2161
/label=rrnB T2 terminator
/note="transcription terminator T2 from the E. coli rrnB
gene"
terminator 2253..2339
/gene="Escherichia coli rrnB"
/label=rrnB T1 terminator
/note="transcription terminator T1 from the E. coli rrnB
gene"
rep_origin complement(2504..3092)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
primer_bind complement(2584..2603)
/label=pBR322ori-F
/note="pBR322 origin, forward primer"
CDS complement(join(3317..3656,1..320))
/codon_start=1
/gene="cat"
/product="chloramphenicol acetyltransferase"
/label=CmR
/note="confers resistance to chloramphenicol"
/translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
KTVKKSKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"