pHR-SFFV-KRAB-dCas9-P2A-mCherry

Lentiviral CRISPR interference vector for expressing mCherry together with a KRAB domain fused to catalytically inactive dCas9.

Sequence Author: Weissman Lab / Addgene #60954

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SV40 NLS SV40 NLS ATG cPPT/CTS pHR-SFFV-KRAB-dCas9-P2A-mCherry 14,247 bp
ATG
8354 .. 8356  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
8354 .. 8356  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
KRAB
8384 .. 8569  =  186 bp
62 amino acids  =  7.4 kDa
Product: Krüppel-associated box (KRAB) transcriptional repression domain from the human zinc finger protein ZNF10 (Margolin et al., 1994)
KRAB
8384 .. 8569  =  186 bp
62 amino acids  =  7.4 kDa
Product: Krüppel-associated box (KRAB) transcriptional repression domain from the human zinc finger protein ZNF10 (Margolin et al., 1994)
dCas9
8591 .. 12,694  =  4104 bp
1368 amino acids  =  158.3 kDa
5 segments
   Segment 1:  
   8591 .. 8617  =  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
8591 .. 12,694  =  4104 bp
1368 amino acids  =  158.3 kDa
5 segments
   Segment 2:  
   8618 .. 8620  =  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
8591 .. 12,694  =  4104 bp
1368 amino acids  =  158.3 kDa
5 segments
   Segment 3:  
   8621 .. 11,107  =  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
8591 .. 12,694  =  4104 bp
1368 amino acids  =  158.3 kDa
5 segments
   Segment 4:  
   11,108 .. 11,110  =  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
8591 .. 12,694  =  4104 bp
1368 amino acids  =  158.3 kDa
5 segments
   Segment 5:  
   11,111 .. 12,694  =  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
8591 .. 12,694  =  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
HA
12,698 .. 12,724  =  27 bp
9 amino acids  =  1.1 kDa
Product: HA (human influenza hemagglutinin) epitope tag
HA
12,698 .. 12,724  =  27 bp
9 amino acids  =  1.1 kDa
Product: HA (human influenza hemagglutinin) epitope tag
SV40 NLS
12,743 .. 12,763  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 NLS
12,743 .. 12,763  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 NLS
12,770 .. 12,790  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 NLS
12,770 .. 12,790  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
P2A
12,830 .. 12,886  =  57 bp
19 amino acids  =  2.0 kDa
Product: 2A peptide from porcine teschovirus-1 polyprotein
Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides.
P2A
12,830 .. 12,886  =  57 bp
19 amino acids  =  2.0 kDa
Product: 2A peptide from porcine teschovirus-1 polyprotein
Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides.
mCherry
12,887 .. 13,597  =  711 bp
236 amino acids  =  26.7 kDa
Product: monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)
mammalian codon-optimized
mCherry
12,887 .. 13,597  =  711 bp
236 amino acids  =  26.7 kDa
Product: monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)
mammalian codon-optimized
AmpR
1277 .. 2137  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   1277 .. 1345  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
1277 .. 2137  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   1346 .. 2137  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
1277 .. 2137  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
LTR
5548 .. 6181  =  634 bp
long terminal repeat (LTR) from HIV-1
LTR
5548 .. 6181  =  634 bp
long terminal repeat (LTR) from HIV-1
ori
2308 .. 2896  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2308 .. 2896  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
WPRE
13,657 .. 14,245  =  589 bp
woodchuck hepatitis virus posttranscriptional regulatory element
WPRE
13,657 .. 14,245  =  589 bp
woodchuck hepatitis virus posttranscriptional regulatory element
SFFV promoter
7866 .. 8273  =  408 bp
spleen focus-forming virus long terminal repeat (LTR) promoter
SFFV promoter
7866 .. 8273  =  408 bp
spleen focus-forming virus long terminal repeat (LTR) promoter
SV40 promoter
3142 .. 3471  =  330 bp
SV40 enhancer and early promoter
SV40 promoter
3142 .. 3471  =  330 bp
SV40 enhancer and early promoter
LTR (ΔU3)
108 .. 341  =  234 bp
self-inactivating long terminal repeat (LTR) from HIV-1
LTR (ΔU3)
108 .. 341  =  234 bp
self-inactivating long terminal repeat (LTR) from HIV-1
RRE
6850 .. 7082  =  233 bp
The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm.
RRE
6850 .. 7082  =  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
5245 .. 5379  =  135 bp
SV40 polyadenylation signal
SV40 poly(A) signal
5245 .. 5379  =  135 bp
SV40 polyadenylation signal
HIV-1 Ψ
6228 .. 6353  =  126 bp
packaging signal of human immunodeficiency virus type 1
HIV-1 Ψ
6228 .. 6353  =  126 bp
packaging signal of human immunodeficiency virus type 1
cPPT/CTS
7596 .. 7713  =  118 bp
central polypurine tract and central termination sequence of HIV-1
cPPT/CTS
7596 .. 7713  =  118 bp
central polypurine tract and central termination sequence of HIV-1
AmpR promoter
1172 .. 1276  =  105 bp
AmpR promoter
1172 .. 1276  =  105 bp
SV40 ori
3322 .. 3457  =  136 bp
SV40 origin of replication
SV40 ori
3322 .. 3457  =  136 bp
SV40 origin of replication
ORF:  574 .. 921  =  348 bp
ORF:  115 amino acids  =  11.9 kDa
ORF:  3754 .. 4128  =  375 bp
ORF:  124 amino acids  =  14.0 kDa
ORF:  7189 .. 7788  =  600 bp
ORF:  199 amino acids  =  23.6 kDa
ORF:  13,747 .. 27  =  528 bp
ORF:  175 amino acids  =  19.3 kDa
ORF:  1277 .. 2137  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  3908 .. 4189  =  282 bp
ORF:  93 amino acids  =  10.7 kDa
ORF:  4220 .. 4603  =  384 bp
ORF:  127 amino acids  =  14.9 kDa
ORF:  6728 .. 7066  =  339 bp
ORF:  112 amino acids  =  12.4 kDa
ORF:  7886 .. 8137  =  252 bp
ORF:  83 amino acids  =  9.2 kDa
ORF:  8354 .. 13,597  =  5244 bp
ORF:  1747 amino acids  =  200.6 kDa
ORF:  13,760 .. 187  =  675 bp
ORF:  224 amino acids  =  24.0 kDa
ORF:  4749 .. 5222  =  474 bp
ORF:  157 amino acids  =  18.2 kDa
ORF:  490 .. 801  =  312 bp
ORF:  103 amino acids  =  10.7 kDa
ORF:  1741 .. 2007  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  3802 .. 4221  =  420 bp
ORF:  139 amino acids  =  15.9 kDa
ORF:  6562 .. 7023  =  462 bp
ORF:  153 amino acids  =  16.6 kDa
ORF:  9940 .. 10,233  =  294 bp
ORF:  97 amino acids  =  10.7 kDa
ORF:  10,504 .. 10,953  =  450 bp
ORF:  149 amino acids  =  17.3 kDa
ORF:  12,718 .. 12,954  =  237 bp
ORF:  78 amino acids  =  8.5 kDa
ORF:  7694 .. 8101  =  408 bp
ORF:  135 amino acids  =  15.2 kDa
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