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Plasmid Files

PX551

Zhang lab AAV vector, also known as AAV-SpCas9 or pAAV-SpCas9, for expressing Cas9 in neurons. Use together with PX552.

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PX551.dna
Map and Sequence File:    Download    Open   
Sequence Author:  Zhang Lab / Addgene #60957
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XbaI (148) AflIII - PciI (7395) SacII (6442) BpmI (6438) AseI (6332) FspI (6284) PvuI (6138) PsiI (5236) BtgZI (5103) PluTI (4808) SfoI (4806) NarI (4805) KasI (4804) NotI (4656) EcoRI (4601) BsmI (4101) BsaBI * (3959) SphI (3727) HindIII (154) MfeI (212) AarI (300) AgeI - SgrAI (389) PspOMI (1816) ApaI (1820) PasI (2230) EcoRV (2314) Eco53kI (2698) SacI (2700) PmlI (3250) PX551 7400 bp
XbaI  (148)
1 site
T C T A G A A G A T C T
AflIII  (7395)
1 site
A C R Y G T T G Y R C A

Sticky ends from different AflIII sites may not be compatible.
PciI  (7395)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
SacII  (6442)
1 site
C C G C G G G G C G C C

Efficient cleavage requires at least two copies of the SacII recognition sequence.
BpmI  (6438)
1 site
C T G G A G ( N ) 14 N N G A C C T C ( N ) 14

Efficient cleavage requires at least two copies of the BpmI recognition sequence.
Sticky ends from different BpmI sites may not be compatible.
After cleavage, BpmI can remain bound to DNA and alter its electrophoretic mobility.
BpmI quickly loses activity at 37°C.
AseI  (6332)
1 site
A T T A A T T A A T T A
FspI  (6284)
1 site
T G C G C A A C G C G T
PvuI  (6138)
1 site
C G A T C G G C T A G C
PsiI  (5236)
1 site
T T A T A A A A T A T T
BtgZI  (5103)
1 site
G C G A T G ( N ) 10 C G C T A C ( N ) 10 ( N ) 4

Sticky ends from different BtgZI sites may not be compatible.
After cleavage, BtgZI can remain bound to DNA and alter its electrophoretic mobility.
BtgZI is typically used at 60°C, but is 75% active at 37°C.
PluTI  (4808)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the PluTI recognition sequence.
SfoI  (4806)
1 site
G G C G C C C C G C G G
NarI  (4805)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the NarI recognition sequence.
KasI  (4804)
1 site
G G C G C C C C G C G G
NotI  (4656)
1 site
G C G G C C G C C G C C G G C G
EcoRI  (4601)
1 site
G A A T T C C T T A A G
BsmI  (4101)
1 site
G A A T G C N C T T A C G N

Sticky ends from different BsmI sites may not be compatible.
BsaBI  (3959)
1 site
G A T N N N N A T C C T A N N N N T A G
* Blocked by Dam methylation.
SphI  (3727)
1 site
G C A T G C C G T A C G
HindIII  (154)
1 site
A A G C T T T T C G A A
MfeI  (212)
1 site
C A A T T G G T T A A C
AarI  (300)
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.
AgeI  (389)
1 site
A C C G G T T G G C C A
SgrAI  (389)
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.
PspOMI  (1816)
1 site
G G G C C C C C C G G G
ApaI  (1820)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PasI  (2230)
1 site
C C C W G G G G G G W C C C

Sticky ends from different PasI sites may not be compatible.
EcoRV  (2314)
1 site
G A T A T C C T A T A G

EcoRV is reportedly more prone than its isoschizomer Eco32I to delete a base after cleavage.
Eco53kI  (2698)
1 site
G A G C T C C T C G A G
SacI  (2700)
1 site
G A G C T C C T C G A G
PmlI  (3250)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
Cas9
464 .. 4564  =  4101 bp
1367 amino acids  =  158.3 kDa
Product: Cas9 (Csn1) endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
generates RNA-guided double strand breaks in DNA
Cas9
464 .. 4564  =  4101 bp
1367 amino acids  =  158.3 kDa
Product: Cas9 (Csn1) endonuclease from the Streptococcus pyogenes Type II CRISPR/Cas system
generates RNA-guided double strand breaks in DNA
AmpR
5720 .. 6580  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   5720 .. 5788  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
5720 .. 6580  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   5789 .. 6580  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
5720 .. 6580  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
6751 .. 7339  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
6751 .. 7339  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
4878 .. 5333  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
4878 .. 5333  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
Mecp2 promoter
160 .. 388  =  229 bp
promoter for selective expression in neurons (Adachi et al., 2005; Gray et al., 2011)
Mecp2 promoter
160 .. 388  =  229 bp
promoter for selective expression in neurons (Adachi et al., 2005; Gray et al., 2011)
AAV2 ITR
1 .. 141  =  141 bp
inverted terminal repeat of adeno-associated virus serotype 2
AAV2 ITR
1 .. 141  =  141 bp
inverted terminal repeat of adeno-associated virus serotype 2
AAV2 ITR
4663 .. 4803  =  141 bp
inverted terminal repeat of adeno-associated virus serotype 2
AAV2 ITR
4663 .. 4803  =  141 bp
inverted terminal repeat of adeno-associated virus serotype 2
AmpR promoter
5615 .. 5719  =  105 bp
AmpR promoter
5615 .. 5719  =  105 bp
poly(A) signal
4607 .. 4655  =  49 bp
synthetic polyadenylation signal
poly(A) signal
4607 .. 4655  =  49 bp
synthetic polyadenylation signal
HA
404 .. 430  =  27 bp
9 amino acids  =  1.1 kDa
Product: HA (human influenza hemagglutinin) epitope tag
HA
404 .. 430  =  27 bp
9 amino acids  =  1.1 kDa
Product: HA (human influenza hemagglutinin) epitope tag
SV40 NLS
434 .. 454  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 NLS
434 .. 454  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 NLS
4568 .. 4588  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
SV40 NLS
4568 .. 4588  =  21 bp
7 amino acids  =  883.1 Da
Product: nuclear localization signal of SV40 large T antigen
ATG
401 .. 403  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
401 .. 403  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ORF:  401 .. 4600  =  4200 bp
ORF:  1399 amino acids  =  162.0 kDa
ORF:  5720 .. 6580  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  264 .. 521  =  258 bp
ORF:  85 amino acids  =  9.3 kDa
ORF:  4677 .. 4901  =  225 bp
ORF:  74 amino acids  =  8.3 kDa
ORF:  84 .. 410  =  327 bp
ORF:  108 amino acids  =  11.5 kDa
ORF:  230 .. 2980  =  2751 bp
ORF:  916 amino acids  =  96.4 kDa
ORF:  4628 .. 4858  =  231 bp
ORF:  76 amino acids  =  8.8 kDa
ORF:  1810 .. 2103  =  294 bp
ORF:  97 amino acids  =  10.8 kDa
ORF:  2374 .. 2823  =  450 bp
ORF:  149 amino acids  =  17.3 kDa
ORF:  6184 .. 6450  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
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