pET28a Cas9-Cys

Bacterial plasmid for regulated expression of S. pyogenes Cas9 with a C-terminal cysteine.

Sequence Author: Kim Lab / Addgene #53261

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Cys SV40 NLS T7 promoter lac operator ATG thrombin site pET28a/Cas9-Cys 9546 bp
ATG
2802 .. 2804  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
2802 .. 2804  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
6xHis
2814 .. 2831  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
2814 .. 2831  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
thrombin site
2841 .. 2858  =  18 bp
6 amino acids  =  627.7 Da
Product: thrombin recognition and cleavage site
thrombin site
2841 .. 2858  =  18 bp
6 amino acids  =  627.7 Da
Product: thrombin recognition and cleavage site
T7 tag (gene 10 leader)
2862 .. 2894  =  33 bp
11 amino acids  =  1.1 kDa
Product: leader peptide from bacteriophage T7 gene 10
promotes efficient translation in E. coli
T7 tag (gene 10 leader)
2862 .. 2894  =  33 bp
11 amino acids  =  1.1 kDa
Product: leader peptide from bacteriophage T7 gene 10
promotes efficient translation in E. coli
Cas9
2937 .. 7040  =  4104 bp
1368 amino acids  =  158.4 kDa
Product: Cas9 (Csn1) endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
generates RNA-guided double strand breaks in DNA
Cas9
2937 .. 7040  =  4104 bp
1368 amino acids  =  158.4 kDa
Product: Cas9 (Csn1) endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
generates RNA-guided double strand breaks in DNA
SV40 NLS
7056 .. 7076  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 NLS
7056 .. 7076  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
HA
7077 .. 7103  =  27 bp
9 amino acids  =  1.1 kDa
Product: HA (human influenza hemagglutinin) epitope tag
HA
7077 .. 7103  =  27 bp
9 amino acids  =  1.1 kDa
Product: HA (human influenza hemagglutinin) epitope tag
Cys
7104 .. 7106  =  3 bp
1 amino acid  =  121.2 Da
Cys
7104 .. 7106  =  3 bp
1 amino acid  =  121.2 Da
lacI
1246 .. 2328  =  1083 bp
360 amino acids  =  38.6 kDa
Product: lac repressor
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
lacI
1246 .. 2328  =  1083 bp
360 amino acids  =  38.6 kDa
Product: lac repressor
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
KanR
7840 .. 8655  =  816 bp
271 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418 (Geneticin®) in eukaryotes
KanR
7840 .. 8655  =  816 bp
271 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418 (Geneticin®) in eukaryotes
ori
8777 .. 9365  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
8777 .. 9365  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
7293 .. 7748  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
7293 .. 7748  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
rop
246 .. 437  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein, which maintains plasmids at low copy number
rop
246 .. 437  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein, which maintains plasmids at low copy number
lacI promoter
2329 .. 2406  =  78 bp
lacI promoter
2329 .. 2406  =  78 bp
T7 terminator
7209 .. 7256  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
T7 terminator
7209 .. 7256  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
lac operator
2734 .. 2758  =  25 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
lac operator
2734 .. 2758  =  25 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
RBS
2773 .. 2795  =  23 bp
efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)
RBS
2773 .. 2795  =  23 bp
efficient ribosome binding site from bacteriophage T7 gene 10 (Olins and Rangwala, 1989)
T7 promoter
2715 .. 2733  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
2715 .. 2733  =  19 bp
promoter for bacteriophage T7 RNA polymerase
ORF:  469 .. 837  =  369 bp
ORF:  122 amino acids  =  14.2 kDa
ORF:  2504 .. 2743  =  240 bp
ORF:  79 amino acids  =  8.0 kDa
ORF:  1212 .. 1463  =  252 bp
ORF:  83 amino acids  =  9.1 kDa
ORF:  2367 .. 2630  =  264 bp
ORF:  87 amino acids  =  9.5 kDa
ORF:  2802 .. 7109  =  4308 bp
ORF:  1435 amino acids  =  165.7 kDa
ORF:  1246 .. 2205  =  960 bp
ORF:  319 amino acids  =  34.1 kDa
ORF:  7840 .. 8655  =  816 bp
ORF:  271 amino acids  =  31.0 kDa
ORF:  246 .. 470  =  225 bp
ORF:  74 amino acids  =  8.5 kDa
ORF:  834 .. 1190  =  357 bp
ORF:  118 amino acids  =  13.0 kDa
ORF:  1226 .. 1489  =  264 bp
ORF:  87 amino acids  =  8.9 kDa
ORF:  2078 .. 2578  =  501 bp
ORF:  166 amino acids  =  17.5 kDa
ORF:  2828 .. 3064  =  237 bp
ORF:  78 amino acids  =  8.8 kDa
ORF:  4286 .. 4579  =  294 bp
ORF:  97 amino acids  =  10.9 kDa
ORF:  4901 .. 5299  =  399 bp
ORF:  132 amino acids  =  15.6 kDa
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