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Plasmid Files

pCas

Bacterial plasmid with a temperature-sensitive replicon encoding Cas9, an inducible single guide RNA (sgRNA), and the λ Red recombination system.

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pCas.dna
Map and Sequence File:    Download    Open   
Sequence Author:  Yang Lab / Addgene #62225
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SgrAI (11,788) BglI (11,291) BsrGI (11,117) SalI (10,868) BspQI - SapI (10,120) AarI (9147) XbaI (9054) PmlI (7209) KpnI (6694) Acc65I (6690) BglII (0) PspOMI (924) ApaI (928) SnaBI (1879) SpeI (2715) EagI - NotI (3035) AsiSI - PvuI (3957) EcoNI (4042) PaeR7I - PspXI - XhoI (4353) SbfI (4504) BstZ17I (5131) BmgBI (5136) SwaI (5437) BlpI (5596) NheI (6015) BmtI (6019) BstBI (6254) pCas 12,545 bp
SgrAI  (11,788)
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.
BglI  (11,291)
1 site
G C C N N N N N G G C C G G N N N N N C C G

Sticky ends from different BglI sites may not be compatible.
BsrGI  (11,117)
1 site
T G T A C A A C A T G T

BsrGI is typically used at 37°C, but is even more active at 60°C.
SalI  (10,868)
1 site
G T C G A C C A G C T G
BspQI  (10,120)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different BspQI sites may not be compatible.
SapI  (10,120)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different SapI sites may not be compatible.
SapI gradually settles in solution, so a tube of SapI should be mixed before removing an aliquot.
AarI  (9147)
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.
XbaI  (9054)
1 site
T C T A G A A G A T C T
PmlI  (7209)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
KpnI  (6694)
1 site
G G T A C C C C A T G G
Acc65I  (6690)
1 site
G G T A C C C C A T G G
BglII  (0)
1 site
A G A T C T T C T A G A
PspOMI  (924)
1 site
G G G C C C C C C G G G
ApaI  (928)
1 site
G G