pMCSG19C

Bacterial expression vector with an MBP-TVMV-6xHis-TEV leader and inducible TVMV protease. See also pMCSG19B.

Sequence Author: Midwest Center for Structural Genomics

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6xHis ScaI (6773) AhdI (6293) PciI (5400) BspQI - SapI (5284) BstZ17I (5171) PflFI - Tth111I (5145) Bpu10I (4506) PpuMI (4406) FspAI (4381) PshAI (4144) HpaI (3805) EcoRV (3749) EagI - NotI (166) SalI (179) Eco53kI (188) SacI (190) BamHI (198) SspI (219) TEV site Acc65I (244) KpnI (248) 6xHis TVMV site BsrGI (321) BsmI (824) BmgBI (900) BglII (1079) PsiI (1090) BsiWI (1148) RBS T7 promoter BspDI * - ClaI * (1549) SgrAI (1591) SphI (1747) ZraI (1751) AatII (1753) PLtetO-1 promoter BseRI (1989) NsiI (2111) AgeI (2154) TVMV protease DraIII (2322) SpeI (2391) SacII (2510) SphI (2774) EcoNI (2834) PflMI (2881) BstEII (3480) PspOMI (3506) ApaI (3510) pMCSG19C 7468 bp
ScaI  (6773)
1 site
A G T A C T T C A T G A
AhdI  (6293)
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.
PciI  (5400)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspQI  (5284)
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  (5284)
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.
BstZ17I  (5171)
1 site
G T A T A C C A T A T G
PflFI  (5145)
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  (5145)
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.
Bpu10I  (4506)
1 site
C C T N A G C G G A N T C G

Cleavage may be enhanced when more than one copy of the Bpu10I recognition sequence is present.
This recognition sequence is asymmetric, so ligating sticky ends generated by Bpu10I will not always regenerate a Bpu10I site.
Sticky ends from different Bpu10I sites may not be compatible.
PpuMI  (4406)
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.
FspAI  (4381)
1 site
R T G C G C A Y Y A C G C G T R
PshAI  (4144)
1 site
G A C N N N N G T C C T G N N N N C A G

PshAI quickly loses activity at 37°C, but can be used at 25°C for long incubations.
HpaI  (3805)
1 site
G T T A A C C A A T T G
EcoRV  (3749)
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.
EagI  (166)
1 site
C G G C C G G C C G G C
NotI  (166)
1 site
G C G G C C G C C G C C G G C G
SalI  (179)
1 site
G T C G A C C A G C T G
Eco53kI  (188)
1 site
G A G C T C C T C G A G
SacI  (190)
1 site
G A G C T C C T C G A G
BamHI  (198)
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.
SspI  (219)
1 site
A A T A T T T T A T A A
Acc65I  (244)
1 site
G G T A C C C C A T G G
KpnI  (248)
1 site
G G T A C C C C A T G G
BsrGI  (321)
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.
BsmI  (824)
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.
BmgBI  (900)
1 site
C A C G T C G T G C A G

This recognition sequence is asymmetric, so ligating blunt ends generated by BmgBI will not always regenerate a BmgBI site.
BglII  (1079)
1 site
A G A T C T T C T A G A
PsiI  (1090)
1 site
T T A T A A A A T A T T
BsiWI  (1148)
1 site
C G T A C G G C A T G C

BsiWI is typically used at 55°C, but is 50% active at 37°C.
BspDI  (1549)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (1549)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
SgrAI  (1591)
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.
SphI  (1747)
2 sites
G C A T G C C G T A C G
ZraI  (1751)
1 site
G A