pLV hUbC-dCas9 VP64-T2A-GFP

Lentiviral vector for expressing enhanced GFP (EGFP) together with catalytically inactive dCas9 fused to the VP64 transcriptional activation domain.

Sequence Author: Gersbach Lab / Addgene #53192

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AmpR promoter SV40 NLS HIV-1 Ψ ATG SV40 NLS pLV hUbC-dCas9 VP64-T2A-GFP 14,937 bp
ATG
4369 .. 4371  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
4369 .. 4371  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
3xFLAG
4372 .. 4437  =  66 bp
22 amino acids  =  2.7 kDa
Product: three tandem FLAG® epitope tags, followed by an enterokinase cleavage site
3xFLAG
4372 .. 4437  =  66 bp
22 amino acids  =  2.7 kDa
Product: three tandem FLAG® epitope tags, followed by an enterokinase cleavage site
SV40 NLS
4444 .. 4464  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 NLS
4444 .. 4464  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
dCas9
4474 .. 8577  =  4104 bp
1368 amino acids  =  158.3 kDa
5 segments
   Segment 1:  
   4474 .. 4500  =  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
4474 .. 8577  =  4104 bp
1368 amino acids  =  158.3 kDa
5 segments
   Segment 2:  
   4501 .. 4503  =  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
4474 .. 8577  =  4104 bp
1368 amino acids  =  158.3 kDa
5 segments
   Segment 3:  
   4504 .. 6990  =  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
4474 .. 8577  =  4104 bp
1368 amino acids  =  158.3 kDa
5 segments
   Segment 4:  
   6991 .. 6993  =  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
4474 .. 8577  =  4104 bp
1368 amino acids  =  158.3 kDa
5 segments
   Segment 5:  
   6994 .. 8577  =  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
4474 .. 8577  =  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
SV40 NLS
8590 .. 8610  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 NLS
8590 .. 8610  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
VP64
8623 .. 8772  =  150 bp
50 amino acids  =  5.5 kDa
Product: tetrameric repeat of the minimal activation domain of herpes simplex virus VP16 (Beerli et al., 1998)
VP64
8623 .. 8772  =  150 bp
50 amino acids  =  5.5 kDa
Product: tetrameric repeat of the minimal activation domain of herpes simplex virus VP16 (Beerli et al., 1998)
T2A
8782 .. 8835  =  54 bp
18 amino acids  =  1.8 kDa
Product: 2A peptide from Thosea asigna virus capsid protein
Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides.
T2A
8782 .. 8835  =  54 bp
18 amino acids  =  1.8 kDa
Product: 2A peptide from Thosea asigna virus capsid protein
Eukaryotic ribosomes fail to insert a peptide bond between the Gly and Pro residues, yielding separate polypeptides.
EGFP
8842 .. 9555  =  714 bp
238 amino acids  =  26.8 kDa
2 segments
   Segment 1:  1a  
   8842 .. 8844  =  3 bp
   1 amino acid  =  117.1 Da
Product: enhanced GFP
mammalian codon-optimized
EGFP
8842 .. 9555  =  714 bp
238 amino acids  =  26.8 kDa
2 segments
   Segment 2:  
   8845 .. 9555  =  711 bp
   237 amino acids  =  26.7 kDa
Product: enhanced GFP
mammalian codon-optimized
EGFP
8842 .. 9555  =  714 bp
238 amino acids  =  26.8 kDa
2 segments
Product: enhanced GFP
mammalian codon-optimized
UbC promoter
3124 .. 4334  =  1211 bp
human ubiquitin C promoter
UbC promoter
3124 .. 4334  =  1211 bp
human ubiquitin C promoter
AmpR
13,944 .. 14,804  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   13,944 .. 14,735  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
13,944 .. 14,804  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   14,736 .. 14,804  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
13,944 .. 14,804  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
13,185 .. 13,773  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
13,185 .. 13,773  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
WPRE
9629 .. 10,217  =  589 bp
woodchuck hepatitis virus posttranscriptional regulatory element
WPRE
9629 .. 10,217  =  589 bp
woodchuck hepatitis virus posttranscriptional regulatory element
f1 ori
11,224 .. 11,652  =  429 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
11,224 .. 11,652  =  429 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
CMV enhancer
238 .. 617  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
238 .. 617  =  380 bp
human cytomegalovirus immediate early enhancer
BleoR
12,109 .. 12,483  =  375 bp
124 amino acids  =  13.8 kDa
Product: antibiotic-binding protein
confers resistance to bleomycin, phleomycin, and Zeocin™
BleoR
12,109 .. 12,483  =  375 bp
124 amino acids  =  13.8 kDa
Product: antibiotic-binding protein
confers resistance to bleomycin, phleomycin, and Zeocin™
SV40 promoter
11,666 .. 11,995  =  330 bp
SV40 enhancer and early promoter
SV40 promoter
11,666 .. 11,995  =  330 bp
SV40 enhancer and early promoter
RRE
1680 .. 1913  =  234 bp
The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm.
RRE
1680 .. 1913  =  234 bp
The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm.
bGH poly(A) signal
10,954 .. 11,178  =  225 bp
bovine growth hormone polyadenylation signal
bGH poly(A) signal
10,954 .. 11,178  =  225 bp
bovine growth hormone polyadenylation signal
CMV promoter
619 .. 817  =  199 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
619 .. 817  =  199 bp
human cytomegalovirus (CMV) immediate early promoter
LTR (truncated)
10,742 .. 10,922  =  181 bp
truncated long terminal repeat (LTR) from HIV-1
LTR (truncated)
10,742 .. 10,922  =  181 bp
truncated long terminal repeat (LTR) from HIV-1
LTR (truncated)
835 .. 1015  =  181 bp
truncated long terminal repeat (LTR) from HIV-1
LTR (truncated)
835 .. 1015  =  181 bp
truncated long terminal repeat (LTR) from HIV-1
HIV-1 Ψ
1062 .. 1187  =  126 bp
packaging signal of human immunodeficiency virus type 1
HIV-1 Ψ
1062 .. 1187  =  126 bp
packaging signal of human immunodeficiency virus type 1
SV40 poly(A) signal
12,613 .. 12,734  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
12,613 .. 12,734  =  122 bp
SV40 polyadenylation signal
cPPT/CTS
2440 .. 2557  =  118 bp
central polypurine tract and central termination sequence of HIV-1
cPPT/CTS
2440 .. 2557  =  118 bp
central polypurine tract and central termination sequence of HIV-1
AmpR promoter
14,805 .. 14,909  =  105 bp
AmpR promoter
14,805 .. 14,909  =  105 bp
EM7 promoter
12,043 .. 12,090  =  48 bp
synthetic bacterial promoter
EM7 promoter
12,043 .. 12,090  =  48 bp
synthetic bacterial promoter
loxP
2602 .. 2635  =  34 bp
Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT).
loxP
2602 .. 2635  =  34 bp
Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT).
loxP
9571 .. 9604  =  34 bp
Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT).
loxP
9571 .. 9604  =  34 bp
Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT).
SV40 ori
11,846 .. 11,981  =  136 bp
SV40 origin of replication
SV40 ori
11,846 .. 11,981  =  136 bp
SV40 origin of replication
ORF:  1558 .. 2445  =  888 bp
ORF:  295 amino acids  =  33.8 kDa
ORF:  4369 .. 9564  =  5196 bp
ORF:  1731 amino acids  =  198.9 kDa
ORF:  12,109 .. 12,483  =  375 bp
ORF:  124 amino acids  =  13.8 kDa
ORF:  14,074 .. 14,340  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  8540 .. 8773  =  234 bp
ORF:  77 amino acids  =  8.9 kDa
ORF:  9719 .. 10,768  =  1050 bp
ORF:  349 amino acids  =  39.4 kDa
ORF:  9732 .. 10,244  =  513 bp
ORF:  170 amino acids  =  18.0 kDa
ORF:  11,175 .. 11,441  =  267 bp
ORF:  88 amino acids  =  9.3 kDa
ORF:  4300 .. 4953  =  654 bp
ORF:  217 amino acids  =  24.0 kDa
ORF:  5878 .. 6396  =  519 bp
ORF:  172 amino acids  =  20.1 kDa
ORF:  6460 .. 6735  =  276 bp
ORF:  91 amino acids  =  10.3 kDa
ORF:  7987 .. 8520  =  534 bp
ORF:  177 amino acids  =  20.0 kDa
ORF:  8800 .. 9591  =  792 bp
ORF:  263 amino acids  =  26.5 kDa
ORF:  1473 .. 1853  =  381 bp
ORF:  126 amino acids  =  13.3 kDa
ORF:  2622 .. 2948  =  327 bp
ORF:  108 amino acids  =  12.1 kDa
ORF:  8562 .. 8882  =  321 bp
ORF:  106 amino acids  =  11.0 kDa
ORF:  12,102 .. 12,389  =  288 bp
ORF:  95 amino acids  =  10.1 kDa
ORF:  13,944 .. 14,804  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  3515 .. 3748  =  234 bp
ORF:  77 amino acids  =  8.4 kDa
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