pgRNA-humanized

Lentiviral vector for expressing a guide RNA (gRNA) together with a puromycin resistance marker and mCherry.

Sequence Author: Qi Lab / Addgene #44248

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loxP BstXI (325) gRNA scaffold polIII terminator XhoI (432) pgRNA-humanized 8308 bp
BstXI  (325)
1 site
C C A N N N N N N T G G G G T N N N N N N A C C

Sticky ends from different BstXI sites may not be compatible.
XhoI  (432)
1 site
C T C G A G G A G C T C
PuroR
1090 .. 1686  =  597 bp
199 amino acids  =  21.5 kDa
Product: puromycin N-acetyltransferase
confers resistance to puromycin
PuroR
1090 .. 1686  =  597 bp
199 amino acids  =  21.5 kDa
Product: puromycin N-acetyltransferase
confers resistance to puromycin
T2A
1696 .. 1749  =  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
1696 .. 1749  =  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.
mCherry
1762 .. 2472  =  711 bp
236 amino acids  =  26.7 kDa
Product: monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)
mammalian codon-optimized
mCherry
1762 .. 2472  =  711 bp
236 amino acids  =  26.7 kDa
Product: monomeric derivative of DsRed fluorescent protein (Shaner et al., 2004)
mammalian codon-optimized
AmpR
4699 .. 5559  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   4699 .. 5490  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
4699 .. 5559  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   5491 .. 5559  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
4699 .. 5559  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
WPRE
2581 .. 3169  =  589 bp
woodchuck hepatitis virus posttranscriptional regulatory element
WPRE
2581 .. 3169  =  589 bp
woodchuck hepatitis virus posttranscriptional regulatory element
ori
3940 .. 4528  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
3940 .. 4528  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
CMV enhancer
549 .. 852  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
549 .. 852  =  304 bp
human cytomegalovirus immediate early enhancer
RRE
7387 .. 7620  =  234 bp
The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm.
RRE
7387 .. 7620  =  234 bp
The Rev response element (RRE) of HIV-1 allows for Rev-dependent mRNA export from the nucleus to the cytoplasm.
CMV promoter
853 .. 1056  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
853 .. 1056  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
LTR (truncated)
3698 .. 3878  =  181 bp
truncated long terminal repeat (LTR) from HIV-1
LTR (truncated)
3698 .. 3878  =  181 bp
truncated long terminal repeat (LTR) from HIV-1
LTR (truncated)
6536 .. 6716  =  181 bp
truncated long terminal repeat (LTR) from HIV-1
LTR (truncated)
6536 .. 6716  =  181 bp
truncated long terminal repeat (LTR) from HIV-1
HIV-1 Ψ
6763 .. 6888  =  126 bp
packaging signal of human immunodeficiency virus type 1
HIV-1 Ψ
6763 .. 6888  =  126 bp
packaging signal of human immunodeficiency virus type 1
cPPT/CTS
8147 .. 8264  =  118 bp
central polypurine tract and central termination sequence of HIV-1
cPPT/CTS
8147 .. 8264  =  118 bp
central polypurine tract and central termination sequence of HIV-1
AmpR promoter
5560 .. 5664  =  105 bp
AmpR promoter
5560 .. 5664  =  105 bp
U6 promoter
243 .. 327  =  85 bp
RNA polymerase III promoter for mouse U6 snRNA
U6 promoter
243 .. 327  =  85 bp
RNA polymerase III promoter for mouse U6 snRNA
gRNA scaffold
349 .. 424  =  76 bp
guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system
gRNA scaffold
349 .. 424  =  76 bp
guide RNA scaffold for the Streptococcus pyogenes CRISPR/Cas9 system
loxP
209 .. 242  =  34 bp
Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT).
loxP
209 .. 242  =  34 bp
Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT).
loxP
2492 .. 2525  =  34 bp
Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT).
loxP
2492 .. 2525  =  34 bp
Cre-mediated recombination occurs in the 8-bp core sequence (GCATACAT).
polIII terminator
425 .. 430  =  6 bp
RNA polymerase III transcription terminator
polIII terminator
425 .. 430  =  6 bp
RNA polymerase III transcription terminator
ORF:  1090 .. 2472  =  1383 bp
ORF:  460 amino acids  =  50.6 kDa
ORF:  2671 .. 3225  =  555 bp
ORF:  184 amino acids  =  20.5 kDa
ORF:  2684 .. 3724  =  1041 bp
ORF:  346 amino acids  =  38.3 kDa
ORF:  4829 .. 5095  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  7265 .. 8152  =  888 bp
ORF:  295 amino acids  =  33.8 kDa
ORF:  1714 .. 2508  =  795 bp
ORF:  264 amino acids  =  26.6 kDa
ORF:  4699 .. 5559  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  7174 .. 7560  =  387 bp
ORF:  128 amino acids  =  13.4 kDa
ORF:  1599 .. 1829  =  231 bp
ORF:  76 amino acids  =  7.9 kDa
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