3xFLAG-dCas9/pMXs-puro vector (V012351)

Basic Vector Information

Vector Name:
3xFLAG-dCas9/pMXs-puro
Antibiotic Resistance:
Ampicillin
Length:
10003 bp
Type:
CRISPR Plasmids
Replication origin:
ori
Source/Author:
Fujita T, Fujii H.
Copy Number:
High copy number

3xFLAG-dCas9/pMXs-puro vector Vector Map

3xFLAG-dCas9/pMXs-puro10003 bp50010001500200025003000350040004500500055006000650070007500800085009000950010000long terminal repeat from Moloney murine leukemiavirusMMLV Psigag (truncated)pol regionATG3xFLAGdCas9SV40 NLSSV40 promoterPuroRLTRoriAmpRAmpR promoter

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 ℃.

3xFLAG-dCas9/pMXs-puro vector Sequence

Copy Sequence

Download GeneBank File(.gb)

LOCUS       3xFLAG-dCas9_pMX       10003 bp DNA     circular SYN 01-JAN-1980
DEFINITION  Retroviral vector with a puromycin resistance marker, encoding 
            FLAG(R)-tagged dCas9 for engineered ChIP (enChIP) purification of 
            specific genomic regions.
ACCESSION   .
VERSION     .
KEYWORDS    3xFLAG-dCas9 pMXs-puro
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 10003)
  AUTHORS   Fujita T, Fujii H.
  TITLE     Identification of proteins associated with an IFN-gamma-responsive 
            promoter by a retroviral expression system for enChIP using CRISPR.
  JOURNAL   PLoS ONE 2014;9:e103084.
  PUBMED    25051498
REFERENCE   2  (bases 1 to 10003)
  AUTHORS   Fujii Lab / Addgene #51240
  TITLE     Direct Submission
REFERENCE   3  (bases 1 to 10003)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "PLoS ONE"; 
            date: "2014"; volume: "9"; pages: "e103084"
COMMENT     SGRef: number: 2; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..10003
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     LTR             22..615
                     /note="long terminal repeat from Moloney murine leukemia
                     virus"
     misc_feature    678..1035
                     /label=MMLV Psi
                     /note="packaging signal of Moloney murine leukemia virus
                     (MMLV)"
     CDS             1094..1510
                     /label=gag (truncated)
                     /note="truncated Moloney murine leukemia virus (MMLV) gag
                     gene lacking the start codon"
     misc_feature    1520..1894
                     /label=pol region
                     /note="Moloney murine leukemia virus (MMLV) pol region 
                     containing the splice acceptor site"
     CDS             1948..1950
                     /codon_start=1
                     /product="start codon"
                     /label=start codon
                     /note="ATG"
                     /translation="M"
     CDS             1951..2016
                     /label=3xFLAG
                     /note="three tandem FLAG(R) epitope tags, followed by an 
                     enterokinase cleavage site"
     CDS             2020..6123
                     /label=dCas9
                     /note="catalytically dead mutant of the Cas9 endonuclease
                     from the Streptococcus pyogenes Type II CRISPR/Cas system"
     CDS             6136..6156
                     /codon_start=1
                     /product="nuclear localization signal of SV40 large T
                     antigen"
                     /note="SV40 NLS"
                     /translation="PKKKRKV"
     promoter        6343..6672
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     CDS             6682..7278
                     /label=PuroR
                     /note="puromycin N-acetyltransferase"
     LTR             7386..7977
                     /label=LTR
                     /note="long terminal repeat from Moloney murine leukemia
                     virus"
     rep_origin      complement(8278..8866)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(9033..9890)
                     /label=AmpR
                     /note="beta-lactamase"
     promoter        complement(9891..9995)
                     /label=AmpR promoter

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