pCLIP-Cas9(D10a)-Nickase-EFS-tRFP

Lentiviral CRISPR vector encoding a Cas9 nickase mutant and the red fluorescent protein TurboRFP expressed from the EF-1α core promoter.

Sequence Author: transOMIC

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FLAG CMV enhancer HIV-1 Ψ pCLIP-Cas9-Nickase-EFS-tRFP 10,792 bp
Cas9(D10A)
2539 .. 6642  =  4104 bp
1368 amino acids  =  158.4 kDa
3 segments
   Segment 1:  
   2539 .. 2565  =  27 bp
   9 amino acids  =  1.1 kDa
Product: nickase mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
generates RNA-guided single strand nicks in DNA due to the D10A mutation in the RuvC catalytic domain
Cas9(D10A)
2539 .. 6642  =  4104 bp
1368 amino acids  =  158.4 kDa
3 segments
   Segment 2:  
   2566 .. 2568  =  3 bp
   1 amino acid  =  89.1 Da
Product: nickase mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
generates RNA-guided single strand nicks in DNA due to the D10A mutation in the RuvC catalytic domain
Cas9(D10A)
2539 .. 6642  =  4104 bp
1368 amino acids  =  158.4 kDa
3 segments
   Segment 3:  
   2569 .. 6642  =  4074 bp
   1358 amino acids  =  157.3 kDa
Product: nickase mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
generates RNA-guided single strand nicks in DNA due to the D10A mutation in the RuvC catalytic domain
Cas9(D10A)
2539 .. 6642  =  4104 bp
1368 amino acids  =  158.4 kDa
3 segments
Product: nickase mutant of the Cas9 endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
generates RNA-guided single strand nicks in DNA due to the D10A mutation in the RuvC catalytic domain
nucleoplasmin NLS
6643 .. 6690  =  48 bp
16 amino acids  =  1.7 kDa
Product: bipartite nuclear localization signal from nucleoplasmin
nucleoplasmin NLS
6643 .. 6690  =  48 bp
16 amino acids  =  1.7 kDa
Product: bipartite nuclear localization signal from nucleoplasmin
FLAG
6691 .. 6714  =  24 bp
8 amino acids  =  1.0 kDa
Product: FLAG® epitope tag, followed by an enterokinase cleavage site
FLAG
6691 .. 6714  =  24 bp
8 amino acids  =  1.0 kDa
Product: FLAG® epitope tag, followed by an enterokinase cleavage site
P2A
6724 .. 6780  =  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
6724 .. 6780  =  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.
TurboRFP
6781 .. 7476  =  696 bp
231 amino acids  =  26.1 kDa
Product: red fluorescent protein from Entacmaea quadricolor
mammalian codon-optimized
TurboRFP
6781 .. 7476  =  696 bp
231 amino acids  =  26.1 kDa
Product: red fluorescent protein from Entacmaea quadricolor
mammalian codon-optimized
AmpR
9797 .. 10,657  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   9797 .. 10,588  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
9797 .. 10,657  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   10,589 .. 10,657  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
9797 .. 10,657  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
WPRE
7501 .. 8089  =  589 bp
woodchuck hepatitis virus posttranscriptional regulatory element
WPRE
7501 .. 8089  =  589 bp
woodchuck hepatitis virus posttranscriptional regulatory element
ori
9041 .. 9626  =  586 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
9041 .. 9626  =  586 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
CMV enhancer
27 .. 404  =  378 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
27 .. 404  =  378 bp
human cytomegalovirus immediate early enhancer
SV40 promoter
8623 .. 8952  =  330 bp
SV40 enhancer and early promoter
SV40 promoter
8623 .. 8952  =  330 bp
SV40 enhancer and early promoter
5' LTR (truncated)
411 .. 668  =  258 bp
truncated 5' long terminal repeat (LTR) from HIV-1
5' LTR (truncated)
411 .. 668  =  258 bp
truncated 5' long terminal repeat (LTR) from HIV-1
RRE
1333 .. 1566  =  234 bp
The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm.
RRE
1333 .. 1566  =  234 bp
The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm.
3' LTR (ΔU3)
8294 .. 8527  =  234 bp
self-inactivating 3' long terminal repeat (LTR) from HIV-1
3' LTR (ΔU3)
8294 .. 8527  =  234 bp
self-inactivating 3' long terminal repeat (LTR) from HIV-1
EF-1α core promoter
2303 .. 2514  =  212 bp
core promoter for human elongation factor EF-1α
EF-1α core promoter
2303 .. 2514  =  212 bp
core promoter for human elongation factor EF-1α
HIV-1 Ψ
715 .. 840  =  126 bp
packaging signal of human immunodeficiency virus type 1
HIV-1 Ψ
715 .. 840  =  126 bp
packaging signal of human immunodeficiency virus type 1
cPPT/CTS
2093 .. 2210  =  118 bp
central polypurine tract and central termination sequence of HIV-1
cPPT/CTS
2093 .. 2210  =  118 bp
central polypurine tract and central termination sequence of HIV-1
AmpR promoter
10,658 .. 10,762  =  105 bp
AmpR promoter
10,658 .. 10,762  =  105 bp
ORF:  2539 .. 7476  =  4938 bp
ORF:  1645 amino acids  =  189.4 kDa
ORF:  7591 .. 8373  =  783 bp
ORF:  260 amino acids  =  28.8 kDa
ORF:  1211 .. 2098  =  888 bp
ORF:  295 amino acids  =  33.8 kDa
ORF:  7604 .. 8116  =  513 bp
ORF:  170 amino acids  =  18.0 kDa
ORF:  9927 .. 10,193  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  10,554 .. 31  =  270 bp
ORF:  89 amino acids  =  10.4 kDa
ORF:  9797 .. 10,657  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  1126 .. 1506  =  381 bp
ORF:  126 amino acids  =  13.3 kDa
ORF:  2251 .. 5058  =  2808 bp
ORF:  935 amino acids  =  98.0 kDa
ORF:  3888 .. 4181  =  294 bp
ORF:  97 amino acids  =  10.8 kDa
ORF:  4452 .. 4901  =  450 bp
ORF:  149 amino acids  =  17.3 kDa
ORF:  6882 .. 7127  =  246 bp
ORF:  81 amino acids  =  9.1 kDa
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