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Plasmid Files

pST1374-N-NLS-flag-linker-Cas9-H840A

Plasmid for mammalian cell expression of Cas9 with the H840A nickase mutation and with an N-terminal nuclear localization signal followed by a FLAG® tag.

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pST1374-N-NLS-flag-linker-Cas9-H840A.dna
Map and Sequence File:    Download    Open   
Sequence Author:  Huang Lab / Addgene #51129
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SgrDI (9316) FspI (8617) AhdI (8395) BstZ17I (7123) PflFI - Tth111I (6709) MscI (6438) SmaI (6329) TspMI - XmaI (6327) AvrII (6306) StuI (6305) SexAI * (6073) PmeI (5257) AgeI (5227) ApaI (5174) PspOMI (5170) MfeI (161) SpeI (249) SnaBI (590) Eco53kI (816) SacI (818) NheI (895) BmtI (899) NotI (932) XbaI (964) PflMI (1006) RsrII (1259) Bsu36I (1513) PaeR7I - XhoI (2200) BbvCI (2257) BamHI (2402) BmgBI (3573) BstXI (3852) KflI (4440) pST1374-N-NLS-flag-linker-Cas9-H840A 9318 bp
SgrDI  (9316)
1 site
C G T C G A C G G C A G C T G C
FspI  (8617)
1 site
T G C G C A A C G C G T
AhdI  (8395)
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.
BstZ17I  (7123)
1 site
G T A T A C C A T A T G
PflFI  (6709)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by PflFI may be hard to ligate.
Sticky ends from different PflFI sites may not be compatible.
Tth111I  (6709)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by Tth111I may be hard to ligate.
Sticky ends from different Tth111I sites may not be compatible.
MscI  (6438)
1 site
T G G C C A A C C G G T
SmaI  (6329)
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  (6327)
1 site
C C C G G G G G G C C C
XmaI  (6327)
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.
AvrII  (6306)
1 site
C C T A G G G G A T C C
StuI  (6305)
1 site
A G G C C T T C C G G A
SexAI  (6073)
1 site
A C C W G G T T G G W C C A
* Blocked by Dcm methylation.
Sticky ends from different SexAI sites may not be compatible.
PmeI  (5257)
1 site
G T T T A A A C C A A A T T T G
AgeI  (5227)
1 site
A C C G G T T G G C C A
ApaI  (5174)
1 site
G G G C C C C C C