Basic Vector Information
- Vector Name:
- lenti dCAS-VP64_Blast
- Antibiotic Resistance:
- Ampicillin
- Length:
- 14085 bp
- Type:
- CRISPR Plasmids
- Replication origin:
- ori
- Source/Author:
- Konermann S, Brigham MD, Trevino AE, Joung J, Abudayyeh OO, Barcena
- Copy Number:
- High copy number
- Promoter:
- EF-1α
lenti dCAS-VP64_Blast 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 ℃.
lenti dCAS-VP64_Blast vector Sequence
LOCUS lenti_dCAS-VP64_ 14085 bp DNA circular SYN 01-JAN-1980
DEFINITION Zhang lab lentiviral vector encoding a blasticidin resistance marker
plus catalytically inactive dCas9 fused to the VP64 transcriptional
activation domain.
ACCESSION .
VERSION .
KEYWORDS lenti dCAS-VP64_Blast.
SOURCE synthetic DNA construct
ORGANISM synthetic DNA construct
REFERENCE 1 (bases 1 to 14085)
AUTHORS Konermann S, Brigham MD, Trevino AE, Joung J, Abudayyeh OO, Barcena
C, Hsu PD, Habib N, Gootenberg JS, Nishimasu H, Nureki O, Zhang F.
TITLE Genome-scale transcriptional activation by an engineered CRISPR-Cas9
complex.
JOURNAL Nature 2015;517:583-8.
PUBMED 25494202
REFERENCE 2 (bases 1 to 14085)
AUTHORS Zhang Lab / Addgene #61425
TITLE Direct Submission
REFERENCE 3 (bases 1 to 14085)
AUTHORS .
TITLE Direct Submission
COMMENT SGRef: number: 1; type: "Journal Article"; journalName: "Nature";
date: "2015"; volume: "517"; pages: "583-8"
COMMENT SGRef: number: 2; type: "Journal Article"
FEATURES Location/Qualifiers
source 1..14085
/mol_type="other DNA"
/organism="synthetic DNA construct"
enhancer 238..617
/label=CMV enhancer
/note="human cytomegalovirus immediate early enhancer"
promoter 618..820
/label=CMV promoter
/note="human cytomegalovirus (CMV) immediate early
promoter"
LTR 835..1015
/label=5' LTR (truncated)
/note="truncated 5' long terminal repeat (LTR) from HIV-1"
misc_feature 1062..1187
/label=HIV-1 Psi
/note="packaging signal of human immunodeficiency virus
type 1"
misc_feature 1680..1913
/label=RRE
/note="The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to the
cytoplasm."
CDS 2098..2142
/label=gp41 peptide
/note="antigenic peptide corresponding to amino acids 655
to 669 of the HIV envelope protein gp41 (Lutje Hulsik et
al., 2013)"
CDS 2291..2332
/note="Protein Tat from Human immunodeficiency virus type 1
group M subtype B (isolate WMJ22). Accession#: P12509"
misc_feature 2440..2557
/label=cPPT/CTS
/note="central polypurine tract and central termination
sequence of HIV-1"
promoter 2687..3865
/label=EF-1-alpha promoter
/note="strong constitutive promoter for human elongation
factor EF-1-alpha"
CDS 3878..3880
/codon_start=1
/product="start codon"
/label=start codon
/note="ATG"
/translation="M"
CDS 3881..3928
/codon_start=1
/product="bipartite nuclear localization signal from
nucleoplasmin"
/label=bipartite nuclear localization signal from
nucl
/note="nucleoplasmin NLS"
/translation="KRPAATKKAGQAKKKK"
CDS 3929..8029
/label=dCas9
/note="catalytically dead mutant of the Cas9 endonuclease
from the Streptococcus pyogenes Type II CRISPR/Cas system"
CDS 8084..8104
/label=SV40 NLS
/note="nuclear localization signal of SV40 (simian virus
40) large T antigen"
CDS 8129..8278
/label=VP64
/note="tetrameric repeat of the minimal activation domain
of herpes simplex virus VP16 (Beerli et al., 1998)"
CDS 8300..8353
/label=T2A
/note="2A peptide from Thosea asigna virus capsid protein"
CDS 8354..8749
/label=BSD
/note="blasticidin S deaminase"
misc_feature 8777..9365
/label=WPRE
/note="woodchuck hepatitis virus posttranscriptional
regulatory element"
LTR 9890..10070
/label=LTR (truncated)
/note="LTR (truncated)"
/note="truncated long terminal repeat (LTR) from HIV-1"
polyA_signal 10102..10326
/label=bGH poly(A) signal
/note="bovine growth hormone polyadenylation signal"
rep_origin 10372..10800
/label=f1 ori
/note="f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis"
promoter 10814..11143
/label=SV40 promoter
/note="SV40 enhancer and early promoter"
promoter 11191..11238
/label=EM7 promoter
/note="synthetic bacterial promoter"
CDS 11257..11628
/label=BleoR
/note="antibiotic-binding protein"
polyA_signal 11761..11894
/label=SV40 poly(A) signal
/note="SV40 polyadenylation signal"
promoter complement(11979..12009)
/label=lac promoter
/note="promoter for the E. coli lac operon"
protein_bind complement(12024..12045)
/label=CAP binding site
/note="CAP binding activates transcription in the presence
of cAMP."
rep_origin complement(12333..12921)
/direction=LEFT
/label=ori
/note="high-copy-number ColE1/pMB1/pBR322/pUC origin of
replication"
CDS complement(13095..13952)
/label=AmpR
/note="beta-lactamase"
promoter complement(13953..14057)
/label=AmpR promoter
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