Resources
Plasmid Files

pHAGE-TO-nls-st1dCas9-3nls-3XTagBFP2

Lentiviral vector for labeling chromosomal loci by tetracycline-regulatable expression of catalytically dead Streptococcus thermophilus dCas9 fused to three copies of mTagBFP2.

To see this sequence with restriction sites, features, and translations, please download
 SnapGene or the free  SnapGene Viewer.

pHAGE-TO-nls-st1dCas9-3nls-3XTagBFP2 Sequence and MappHAGE-TO-nls-st1dCas9-3nls-3XTagBFP2.dna
Map and Sequence File   
Sequence Author:  Pederson Lab / Addgene #64512
Download Free Trial Get SnapGene Viewer

 AvrII (11,834) SfiI (11,787) PciI (11,555) DrdI (11,453) AhdI (10,667) PvuI (10,298) SspI (9862) PacI (9652) SmaI (9644) TspMI - XmaI (9642) PsiI (9605) XbaI (8667) PaeR7I - XhoI (8653) BspDI - ClaI (7931) NotI (7200) BamHI (6472) SV40 NLS SV40 NLS SV40 NLS SpeI (6382) PmlI (290) NruI * (833) FseI (1150) MfeI (1189) BbvCI (1424) BstAPI (1453) KflI - PpuMI (1934) NheI (2221) BmtI (2225) NdeI (2472) SnaBI (2578) CMV promoter tet operator tet operator SalI - SgrDI (2850) AgeI (2944) MluI (2958) ATG SV40 NLS EcoRI (3007) BstEII (3078) SgrAI (3194) AarI (4215) BstXI (5088) SbfI (5108) AscI (5907) FspAI (6310) pHAGE-TO-nls-st1dCas9-3nls-3XTagBFP2 12,284 bp
AvrII  (11,834)
1 site
C C T A G G G G A T C C
SfiI  (11,787)
1 site
G G C C N N N N N G G C C C C G G N N N N N C C G G

Efficient cleavage requires at least two copies of the SfiI
recognition sequence.
Sticky ends from different SfiI sites may not be compatible.
PciI  (11,555)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
DrdI  (11,453)
1 site
G A C N N N N N N G T C C T G N N N N N N C A G

Sticky ends from different DrdI sites may not be compatible.
AhdI  (10,667)
1 site
G A C N N N N N G T C C T G N N N N N C A G

The 1-base overhangs produced by AhdI may be hard to ligate.
Sticky ends from different AhdI sites may not be compatible.
PvuI  (10,298)
1 site
C G A T C G G C T A G C
SspI  (9862)
1 site
A A T A T T T T A T A A
PacI  (9652)
1 site
T T A A T T A A A A T T A A T T
SmaI  (9644)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
TspMI  (9642)
1 site
C C C G G G G G G C C C
XmaI  (9642)
1 site
C C C G G G G G G C C C

Efficient cleavage requires at least two copies of the XmaI
recognition sequence.
Full cleavage with XmaI may require a long incubation.
PsiI  (9605)
1 site
T T A T A A A A T A T T
XbaI  (8667)
1 site
T C T A G A A G A T C T
PaeR7I  (8653)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
XhoI  (8653)
1 site
C T C G A G G A G C T C
BspDI  (7931)
1 site
A T C G A T T A G C T A
ClaI  (7931)
1 site
A T C G A T T A G C T A
NotI  (7200)
1 site
G C G G C C G C C G C C G G C G
BamHI  (6472)
1 site
G G A T C C C C T A G G

After cleavage, BamHI-HF™ (but not the original BamHI) can
remain bound to DNA and alter its electrophoretic mobility.
SpeI  (6382)
1 site
A C T A G T T G A T C A
PmlI  (290)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
NruI  (833)
1 site
T C G C G A A G C G C T
* Blocked by Dam methylation.
FseI  (1150)
1 site
G G C C G G C C C C G G C C G G

FseI gradually loses activity when stored at -20°C.
MfeI  (1189)
1 site
C A A T T G G T T A A C
BbvCI  (1424)
1 site
C C T C A G C G G A G T C G
BstAPI  (1453)
1 site
G C A N N N N N T G C C G T N N N N N A C G

Sticky ends from different BstAPI sites may not be compatible.
KflI  (1934)
1 site
G G G W C C C C C C W G G G

Sticky ends from different KflI sites may not be compatible.
PpuMI  (1934)
1 site
R G G W C C Y Y C C W G G R

Sticky ends from different PpuMI sites may not be compatible.
NheI  (2221)
1 site
G C T A G C C G A T C G
BmtI  (2225)
1 site
G C T A G C C G A T C G
NdeI  (2472)
1 site
C A T A T G G T A T A C

Prolonged incubation with NdeI may lead to removal of additional
nucleotides.
SnaBI  (2578)
1 site
T A C G T A A T G C A T
SalI  (2850)
1 site
G T C G A C C A G C T G
SgrDI  (2850)
1 site
C G T C G A C G G C A G C T G C
AgeI  (2944)
1 site
A C C G G T T G G C C A

AgeI quickly loses activity at 37°C, but can be used at 25°C for
long incubations.
MluI  (2958)
1 site
A C G C G T T G C G C A
EcoRI  (3007)
1 site
G A A T T C C T T A A G
BstEII  (3078)
1 site
G G T N A C C C C A N T G G

Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
SgrAI  (3194)
1 site
C R C C G G Y G G Y G G C C R C

Efficient cleavage requires at least two copies of the SgrAI
recognition sequence.
AarI  (4215)
1 site
C A C C T G C ( N ) 4 G T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the AarI
recognition sequence.
Sticky ends from different AarI sites may not be compatible.
After cleavage, AarI can remain bound to DNA and alter its
electrophoretic mobility.
BstXI  (5088)
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.
SbfI  (5108)
1 site
C C T G C A G G G G A C G T C C
AscI  (5907)
1 site
G G C G C G C C C C G C G C G G
FspAI  (6310)
1 site
R T G C G C A Y Y A C G C G T R
St1dCas9
3016 .. 6375  =  3360 bp
1120 amino acids  =  129.1 kDa
   Segment 1:  
   3016 .. 3036  =  21 bp
   7 amino acids  =  715.9 Da
Product: catalytically dead mutant of the Cas9
endonuclease from the Streptococcus thermophilus
Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks
endonuclease activity due to the mutations D9A,
D598A, H599A, and N622A
St1dCas9
3016 .. 6375  =  3360 bp
1120 amino acids  =  129.1 kDa
   Segment 2:  
   3037 .. 3039  =  3 bp
   1 amino acid  =  89.1 Da
Product: catalytically dead mutant of the Cas9
endonuclease from the Streptococcus thermophilus
Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks
endonuclease activity due to the mutations D9A,
D598A, H599A, and N622A
St1dCas9
3016 .. 6375  =  3360 bp
1120 amino acids  =  129.1 kDa
   Segment 3:  
   3040 .. 4803  =  1764 bp
   588 amino acids  =  67.5 kDa
Product: catalytically dead mutant of the Cas9
endonuclease from the Streptococcus thermophilus
Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks
endonuclease activity due to the mutations D9A,
D598A, H599A, and N622A
St1dCas9
3016 .. 6375  =  3360 bp
1120 amino acids  =  129.1 kDa
   Segment 4:  
   4804 .. 4809  =  6 bp
   2 amino acids  =  160.2 Da
Product: catalytically dead mutant of the Cas9
endonuclease from the Streptococcus thermophilus
Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks
endonuclease activity due to the mutations D9A,
D598A, H599A, and N622A
St1dCas9
3016 .. 6375  =  3360 bp
1120 amino acids  =  129.1 kDa
   Segment 5:  
   4810 .. 4875  =  66 bp
   22 amino acids  =  2.4 kDa
Product: catalytically dead mutant of the Cas9
endonuclease from the Streptococcus thermophilus
Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks
endonuclease activity due to the mutations D9A,
D598A, H599A, and N622A
St1dCas9
3016 .. 6375  =  3360 bp
1120 amino acids  =  129.1 kDa
   Segment 6:  
   4876 .. 4878  =  3 bp
   1 amino acid  =  89.1 Da
Product: catalytically dead mutant of the Cas9
endonuclease from the Streptococcus thermophilus
Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks
endonuclease activity due to the mutations D9A,
D598A, H599A, and N622A
St1dCas9
3016 .. 6375  =  3360 bp
1120 amino acids  =  129.1 kDa
   Segment 7:  
   4879 .. 6375  =  1497 bp
   499 amino acids  =  58.3 kDa
Product: catalytically dead mutant of the Cas9
endonuclease from the Streptococcus thermophilus
Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks
endonuclease activity due to the mutations D9A,
D598A, H599A, and N622A
St1dCas9
3016 .. 6375  =  3360 bp
1120 amino acids  =  129.1 kDa
7 segments
Product: catalytically dead mutant of the Cas9
endonuclease from the Streptococcus thermophilus
Type II CRISPR/Cas system
RNA-guided DNA-binding protein that lacks
endonuclease activity due to the mutations D9A,
D598A, H599A, and N622A
AmpR
9880 .. 10,740  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 1:  signal sequence  
   9880 .. 9948  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
9880 .. 10,740  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 2:  
   9949 .. 10,740  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
9880 .. 10,740  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
mTagBFP2
6484 .. 7194  =  711 bp
237 amino acids  =  26.7 kDa
Product: enhanced monomeric blue fluorescent
protein (Subach et al., 2011)
mammalian codon-optimized
mTagBFP2
6484 .. 7194  =  711 bp
237 amino acids  =  26.7 kDa
Product: enhanced monomeric blue fluorescent
protein (Subach et al., 2011)
mammalian codon-optimized
mTagBFP2
7213 .. 7923  =  711 bp
237 amino acids  =  26.7 kDa
Product: enhanced monomeric blue fluorescent
protein (Subach et al., 2011)
mammalian codon-optimized
mTagBFP2
7213 .. 7923  =  711 bp
237 amino acids  =  26.7 kDa
Product: enhanced monomeric blue fluorescent
protein (Subach et al., 2011)
mammalian codon-optimized
mTagBFP2
7942 .. 8652  =  711 bp
237 amino acids  =  26.7 kDa
Product: enhanced monomeric blue fluorescent
protein (Subach et al., 2011)
mammalian codon-optimized
mTagBFP2
7942 .. 8652  =  711 bp
237 amino acids  =  26.7 kDa
Product: enhanced monomeric blue fluorescent
protein (Subach et al., 2011)
mammalian codon-optimized
3' LTR
1 .. 634  =  634 bp
3' long terminal repeat (LTR) from HIV-1
3' LTR
1 .. 634  =  634 bp
3' long terminal repeat (LTR) from HIV-1
WPRE
8674 .. 9262  =  589 bp
woodchuck hepatitis virus posttranscriptional
regulatory element
WPRE
8674 .. 9262  =  589 bp
woodchuck hepatitis virus posttranscriptional
regulatory element
ori
10,911 .. 11,499  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
10,911 .. 11,499  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
CMV enhancer
2299 .. 2602  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
2299 .. 2602  =  304 bp
human cytomegalovirus immediate early enhancer
RRE
1303 .. 1536  =  234 bp
The Rev response element (RRE) of HIV-1 allows for
Rev-dependent mRNA export from the nucleus to
the cytoplasm.
RRE
1303 .. 1536  =  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)
9337 .. 9570  =  234 bp
self-inactivating 3' long terminal repeat (LTR) from
HIV-1
3' LTR (ΔU3)
9337 .. 9570  =  234 bp
self-inactivating 3' long terminal repeat (LTR) from
HIV-1
CMV promoter
2603 .. 2806  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
2603 .. 2806  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
HIV-1 Ψ
681 .. 806  =  126 bp
packaging signal of human immunodeficiency virus
type 1
HIV-1 Ψ
681 .. 806  =  126 bp
packaging signal of human immunodeficiency virus
type 1
cPPT/CTS
2058 .. 2175  =  118 bp
central polypurine tract and central termination
sequence of HIV-1
cPPT/CTS
2058 .. 2175  =  118 bp
central polypurine tract and central termination
sequence of HIV-1
AmpR promoter
9775 .. 9879  =  105 bp
AmpR promoter
9775 .. 9879  =  105 bp
SV40 NLS
2986 .. 3006  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T
antigen
SV40 NLS
2986 .. 3006  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T
antigen
SV40 NLS
6391 .. 6411  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T
antigen
SV40 NLS
6391 .. 6411  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T
antigen
SV40 NLS
6424 .. 6444  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T
antigen
SV40 NLS
6424 .. 6444  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T
antigen
SV40 NLS
6451 .. 6471  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T
antigen
SV40 NLS
6451 .. 6471  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T
antigen
tet operator
2808 .. 2826  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
2808 .. 2826  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
2829 .. 2847  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
2829 .. 2847  =  19 bp
bacterial operator O2 for the tetR and tetA genes
ATG
2980 .. 2982  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
2980 .. 2982  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
Try SnapGene and create your own beautiful maps

Individual Sequences & Maps

SnapGene offers the fastest and easiest way to plan, visualize, and document your molecular biology procedures.

Priced accessibly so that everyone in your lab can have a license.

Learn More...

SnapGene Viewer is a versatile tool for creating and sharing richly annotated sequence files. It opens many common file formats.

Free! Because there should be no barriers to seeing your data.

Learn More...

The map, notes, and annotations on this page and in the sequence/map file are copyrighted material. This material may be used without restriction by academic, nonprofit, and governmental entities, except that the source must be cited as "www.snapgene.com/resources". Commercial entities must contact GSL Biotech LLC for permission and terms of use.

Copyright © 2016 GSL Biotech LLC | Site Map | Privacy | Legal Disclaimers   Subscribe to Our Newsletter