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pCBC-DT3T4 vector (V017494)

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 Vector Map

pCBC-DT3T43656 bp60012001800240030003600CAT-RccdBIn lacZ geneT7gRNA scaffoldSP6 promoterM13 revEM7 promoterpENTR-FrrnB T2 terminatorrrnB T1 terminatororiCmR

Plasmid Resuspension 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 ℃.

pCBC-DT3T4 vector Sequence

Copy Sequence

Download GeneBank File(.gb)

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"

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