Resources
Plasmid Files

pHR-SFFV-dCas9-BFP

Lentiviral CRISPR interference vector for expressing catalytically inactive dCas9 fused to TagBFP.

To see this sequence with restriction sites, features, and translations, please download
 SnapGene or the free  SnapGene Viewer.

pHR-SFFV-dCas9-BFP.dna
Map and Sequence File:    Download    Open   
Sequence Author:  Qi & Weissman Labs / Addgene #46910
Download Free Trial Get SnapGene Viewer


HpaI (13,633) AgeI (12,335) AvrII (11,845) SfiI (11,798) FspI (10,230) PvuI (10,084) AfeI (9121) SgrAI (9035) AbsI - PspXI (8403) SbfI (7767) Bsu36I (7290) NruI * (833) FseI (1150) AleI (1576) BstEII (2468) AscI (2692) PflFI - Tth111I (2714) MluI (2754) BamHI (7034) pHR-SFFV-dCas9-BFP 13,936 bp
HpaI  (13,633)
1 site
G T T A A C C A A T T G
AgeI  (12,335)
1 site
A C C G G T T G G C C A
AvrII  (11,845)
1 site
C C T A G G G G A T C C
SfiI  (11,798)
1 site
G G C C N N N N N G G C C C C G G N N N N N C C G G

Efficient cleavage requires at least two copies of the SfiI recognition sequence.
Sticky ends from different SfiI sites may not be compatible.
FspI  (10,230)
1 site
T G C G C A A C G C G T
PvuI  (10,084)
1 site
C G A T C G G C T A G C
AfeI  (9121)
1 site
A G C G C T T C G C G A
SgrAI  (9035)
1 site
C R C C G G Y G G Y G G C C R C

Efficient cleavage requires at least two copies of the SgrAI recognition sequence.
AbsI  (8403)
1 site
C C T C G A G G G G A G C T C C
PspXI  (8403)
1 site
V C T C G A G B B G A G C T C V
SbfI  (7767)
1 site
C C T G C A G G G G A C G T C C
Bsu36I  (7290)
1 site
C C T N A G G G G A N T C C

Sticky ends from different Bsu36I sites may not be compatible.
NruI  (833)
1 site
T C G C G A A G C G C T
* Blocked by Dam methylation.
FseI  (1150)
1 site
G G C C G G C C C C G G C C G G

FseI gradually loses activity when stored at -20°C.
AleI  (1576)
1 site
C A C N N N N G T G G T G N N N N C A C
BstEII  (2468)
1 site
G G T N A C C C C A N T G G

Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
AscI  (2692)
1 site
G G C G C G C C C C G C G C G G
PflFI  (2714)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by PflFI may be hard to ligate.
Sticky ends from different PflFI sites may not be compatible.
Tth111I  (2714)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by Tth111I may be hard to ligate.
Sticky ends from different Tth111I sites may not be compatible.
MluI  (2754)
1 site
A C G C G T T G C G C A
BamHI  (7034)
1 site
G G A T C C C C T A G G

After cleavage, BamHI-HF™ (but not the original BamHI) can remain bound to DNA and alter its electrophoretic mobility.
dCas9
2801 .. 6904  =  4104 bp
1368 amino acids  =  158.3 kDa
   Segment 1:  
   2801 .. 2827  =  27 bp
   9 amino acids  =  1.1 kDa
Product: catalytically dead mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks endonuclease activity due to the D10A mutation in the RuvC catalytic domain and the H840A mutation in the HNH catalytic domain
dCas9
2801 .. 6904  =  4104 bp
1368 amino acids  =  158.3 kDa
   Segment 2:  
   2828 .. 2830  =  3 bp
   1 amino acid  =  89.1 Da
Product: catalytically dead mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks endonuclease activity due to the D10A mutation in the RuvC catalytic domain and the H840A mutation in the HNH catalytic domain
dCas9
2801 .. 6904  =  4104 bp
1368 amino acids  =  158.3 kDa
   Segment 3:  
   2831 .. 5317  =  2487 bp
   829 amino acids  =  96.3 kDa
Product: catalytically dead mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks endonuclease activity due to the D10A mutation in the RuvC catalytic domain and the H840A mutation in the HNH catalytic domain
dCas9
2801 .. 6904  =  4104 bp
1368 amino acids  =  158.3 kDa
   Segment 4:  
   5318 .. 5320  =  3 bp
   1 amino acid  =  89.1 Da
Product: catalytically dead mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks endonuclease activity due to the D10A mutation in the RuvC catalytic domain and the H840A mutation in the HNH catalytic domain
dCas9
2801 .. 6904  =  4104 bp
1368 amino acids  =  158.3 kDa
   Segment 5:  
   5321 .. 6904  =  1584 bp
   528 amino acids  =  60.9 kDa
Product: catalytically dead mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks endonuclease activity due to the D10A mutation in the RuvC catalytic domain and the H840A mutation in the HNH catalytic domain
dCas9
2801 .. 6904  =  4104 bp
1368 amino acids  =  158.3 kDa
5 segments
Product: catalytically dead mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks endonuclease activity due to the D10A mutation in the RuvC catalytic domain and the H840A mutation in the HNH catalytic domain
AmpR
9666 .. 10,526  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   9666 .. 9734  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
9666 .. 10,526  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   9735 .. 10,526  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
9666 .. 10,526  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
TagBFP
7040 .. 7738  =  699 bp
232 amino acids  =  26.2 kDa
Product: monomeric blue fluorescent protein
mammalian codon-optimized
TagBFP
7040 .. 7738  =  699 bp
232 amino acids  =  26.2 kDa
Product: monomeric blue fluorescent protein
mammalian codon-optimized
LTR
1 .. 634  =  634 bp
long terminal repeat (LTR) from HIV-1
LTR
1 .. 634  =  634 bp
long terminal repeat (LTR) from HIV-1
WPRE
7799 .. 8387  =  589 bp
woodchuck hepatitis virus posttranscriptional regulatory element
WPRE
7799 .. 8387  =  589 bp
woodchuck hepatitis virus posttranscriptional regulatory element
ori
10,697 .. 11,285  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
10,697 .. 11,285  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
SFFV promoter
2319 .. 2726  =  408 bp
spleen focus-forming virus long terminal repeat (LTR) promoter
SFFV promoter
2319 .. 2726  =  408 bp
spleen focus-forming virus long terminal repeat (LTR) promoter
SV40 promoter
11,531 .. 11,860  =  330 bp
SV40 enhancer and early promoter
SV40 promoter
11,531 .. 11,860  =  330 bp
SV40 enhancer and early promoter
LTR (ΔU3)
8497 .. 8730  =  234 bp
self-inactivating long terminal repeat (LTR) from HIV-1
LTR (ΔU3)
8497 .. 8730  =  234 bp
self-inactivating long terminal repeat (LTR) from HIV-1
RRE
1303 .. 1535  =  233 bp
The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm.
RRE
1303 .. 1535  =  233 bp
The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm.
SV40 poly(A) signal
13,634 .. 13,768  =  135 bp
SV40 polyadenylation signal
SV40 poly(A) signal
13,634 .. 13,768  =  135 bp
SV40 polyadenylation signal
HIV-1 Ψ
681 .. 806  =  126 bp
packaging signal of human immunodeficiency virus type 1
HIV-1 Ψ
681 .. 806  =  126 bp
packaging signal of human immunodeficiency virus type 1
cPPT/CTS
2049 .. 2166  =  118 bp
central polypurine tract and central termination sequence of HIV-1
cPPT/CTS
2049 .. 2166  =  118 bp
central polypurine tract and central termination sequence of HIV-1
AmpR promoter
9561 .. 9665  =  105 bp
AmpR promoter
9561 .. 9665  =  105 bp
HA
6908 .. 6934  =  27 bp
9 amino acids  =  1.1 kDa
Product: HA (human influenza hemagglutinin) epitope tag
HA
6908 .. 6934  =  27 bp
9 amino acids  =  1.1 kDa
Product: HA (human influenza hemagglutinin) epitope tag
SV40 NLS
6953 .. 6973  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 NLS
6953 .. 6973  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 NLS
6980 .. 7000  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 NLS
6980 .. 7000  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 ori
11,711 .. 11,846  =  136 bp
SV40 origin of replication
SV40 ori
11,711 .. 11,846  =  136 bp
SV40 origin of replication
ORF:  1642 .. 2241  =  600 bp
ORF:  199 amino acids  =  23.6 kDa
ORF:  13,138 .. 13,611  =  474 bp
ORF:  157 amino acids  =  18.2 kDa
ORF:  1181 .. 1519  =  339 bp
ORF:  112 amino acids  =  12.4 kDa
ORF:  2339 .. 2590  =  252 bp
ORF:  83 amino acids  =  9.2 kDa
ORF:  2801 .. 7738  =  4938 bp
ORF:  1645 amino acids  =  189.2 kDa
ORF:  7889 .. 8416  =  528 bp
ORF:  175 amino acids  =  19.3 kDa
ORF:  8963 .. 9310  =  348 bp
ORF:  115 amino acids  =  11.9 kDa
ORF:  12,143 .. 12,517  =  375 bp
ORF:  124 amino acids  =  14.0 kDa
ORF:  7902 .. 8576  =  675 bp
ORF:  224 amino acids  =  24.0 kDa
ORF:  9666 .. 10,526  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  12,297 .. 12,578  =  282 bp
ORF:  93 amino acids  =  10.7 kDa
ORF:  12,609 .. 12,992  =  384 bp
ORF:  127 amino acids  =  14.9 kDa
ORF:  2147 .. 2554  =  408 bp
ORF:  135 amino acids  =  15.2 kDa
ORF:  2675 .. 7543  =  4869 bp
ORF:  1622 amino acids  =  170.5 kDa
ORF:  8879 .. 9190  =  312 bp
ORF:  103 amino acids  =  10.7 kDa
ORF:  10,130 .. 10,396  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  12,191 .. 12,610  =  420 bp
ORF:  139 amino acids  =  15.9 kDa
ORF:  1015 .. 1476  =  462 bp
ORF:  153 amino acids  =  16.6 kDa
ORF:  4150 .. 4443  =  294 bp
ORF:  97 amino acids  =  10.7 kDa
ORF:  4714 .. 5163  =  450 bp
ORF:  149 amino acids  =  17.3 kDa
Try SnapGene and create your own beautiful maps

Individual Sequences & Maps