HBT-pcoCas9 vector (V012338)

MSDS

Basic Vector Information

- - Vector Name:
  - HBT-pcoCas9
- - Antibiotic Resistance:
  - Ampicillin
- - Length:
  - 7940 bp
- - Type:
  - CRISPR Plasmids
- - Replication origin:
  - ori
- - Source/Author:
  - Li JF, Norville JE, Aach J, McCormack M, Zhang D, Bush J, Church GM,
- - Copy Number:
  - High copy number
- - Promoter:
  - 35SPPDK

HBT-pcoCas9 vector Vector Map

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 ℃.

HBT-pcoCas9 vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       HBT-pcoCas9.        7940 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Plasmid for transient expression of plant codon-optimized Cas9 in 
            plant cells. Use together with pUC119-gRNA.
ACCESSION   .
VERSION     .
KEYWORDS    HBT-pcoCas9.
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7940)
  AUTHORS   Li JF, Norville JE, Aach J, McCormack M, Zhang D, Bush J, Church GM,
            Sheen J.
  TITLE     Multiplex and homologous recombination-mediated genome editing in 
            Arabidopsis and Nicotiana benthamiana using guide RNA and Cas9.
  JOURNAL   Nat. Biotechnol. 2013;31:688-91.
  PUBMED    23929339
REFERENCE   2  (bases 1 to 7940)
  AUTHORS   Sheen Lab / Addgene #52254
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 7940)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat. 
            Biotechnol."; date: "2013"; volume: "31"; pages: "688-91"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..7940
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(39..55)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    complement(63..79)
                     /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(87..117)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    complement(132..153)
                     /label=CAP binding site
                     /note="CAP binding activates transcription in the presence
                     of cAMP."
     rep_origin      complement(441..1029)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(1203..2060)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(2061..2165)
                     /label=AmpR promoter
     primer_bind     2639..2655
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        2722..3231
                     /label=35SPPDK promoter
                     /note="hybrid promoter consisting of the cauliflower mosaic
                     virus 35S enhancer fused to the maize C4PPDK basal promoter
                     (Yoo et al., 2007)"
     CDS             3240..3242
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     CDS             3243..3290
                     /label=2xFLAG
                     /note="two tandem FLAG(R) epitope tags"
     CDS             3297..3317
                     /label=SV40 NLS
                     /note="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
     intron          4779..4967
                     /label=IV2 intron
                     /note="modified second intron of the potato ST-LS1 gene 
                     (Vancanneyt et al., 1990)"
     CDS             5133..7631
                     /label=Cas9(C)
                     /note="C-terminal portion of Streptococcus pyogenes Cas9 
                     (Zetsche et al., 2015)"
     CDS             7632..7679
                     /label=nucleoplasmin NLS
                     /note="bipartite nuclear localization signal from
                     nucleoplasmin"
     terminator      7688..7940
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"

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