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pMCSG66

Bacterial vector for expressing a protein with a 6xHis-2xHA-TEV leader plus a second untagged protein.

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pMCSG66.dna
Map and Sequence File:    Download    Open   
Sequence Author:  Midwest Center for Structural Genomics
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ScaI (4701) PvuI (4591) PstI (4466) BsaI (4282) AhdI (4221) AlwNI (3744) PciI (3328) BspQI - SapI (3212) BstZ17I (3099) BsaAI (3080) PflFI - Tth111I (3073) StyI (57) BlpI (80) PaeR7I - PspXI - XhoI (158) EagI - NotI (166) HindIII (173) SalI (179) Eco53kI (188) SacI (190) EcoRI (192) BamHI (198) TspMI - XmaI (232) SmaI (234) SspI (275) Acc65I (300) KpnI (304) BfuAI - BspMI (312) BsiWI (323) BglII (361) NdeI (397) XbaI (435) BspDI * - ClaI * (504) SgrAI (546) SphI (702) EcoNI (762) PflMI (809) BstAPI (910) MluI (1227) BclI * (1241) BstEII (1408) PspOMI (1434) ApaI (1438) BssHII (1638) EcoRV (1677) HpaI (1733) PshAI (2072) FspAI (2309) PpuMI (2334) Bpu10I (2434) pMCSG66 5396 bp
ScaI  (4701)
1 site
A G T A C T T C A T G A
PvuI  (4591)
1 site
C G A T C G G C T A G C
PstI  (4466)
1 site
C T G C A G G A C G T C
BsaI  (4282)
1 site
G G T C T C N C C A G A G N ( N ) 4

Sticky ends from different BsaI sites may not be compatible.
BsaI can be used between 37°C and 50°C.
AhdI  (4221)
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.
AlwNI  (3744)
1 site
C A G N N N C T G G T C N N N G A C

Sticky ends from different AlwNI sites may not be compatible.
PciI  (3328)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspQI  (3212)
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  (3212)
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  (3099)
1 site
G T A T A C C A T A T G
BsaAI  (3080)
1 site
Y A C G T R R T G C A Y
PflFI  (3073)
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  (3073)
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.
StyI  (57)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
BlpI  (80)
1 site
G C T N A G C C G A N T C G

Sticky ends from different BlpI sites may not be compatible.
PaeR7I  (158)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (158)
1 site
V C T C G A G B B G A G C T C V
XhoI  (158)
1 site
C T C G A G G A G C T C
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
HindIII  (173)
1 site
A A G C T T T T C G A A
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
EcoRI  (192)
1 site
G A A T T C C T T A 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.
TspMI  (232)
1 site
C C C G G G G G G C C C
XmaI  (232)
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.
SmaI  (234)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
SspI  (275)
1 site
A A T A T T T T A T A A
Acc65I  (300)
1 site
G G T A C C C C A T G G
KpnI  (304)
1 site
G G T A C C C C A T G G
BfuAI  (312)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BfuAI recognition sequence.
Sticky ends from different BfuAI sites may not be compatible.
BfuAI is typically used at 50°C, but is 50% active at 37°C.
BspMI  (312)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BspMI recognition sequence.
Sticky ends from different BspMI sites may not be compatible.
BsiWI  (323)
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.
BglII  (361)
1 site
A G A T C T T C T A G A
NdeI  (397)
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.
XbaI  (435)
1 site
T C T A G A A G A T C T
BspDI  (504)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (504)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
SgrAI  (546)
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  (702)
1 site
G C A T G C C G T A C G
EcoNI  (762)
1 site
C C T N N N N N A G G G G A N N N N N T C C

The 1-base overhangs produced by EcoNI may be hard to ligate.
Sticky ends from different EcoNI sites may not be compatible.
PflMI  (809)
1 site
C C A N N N N N T G G G G T N N N N N A C C

Sticky ends from different PflMI sites may not be compatible.
BstAPI  (910)
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.
MluI  (1227)
1 site
A C G C G T T G C G C A
BclI  (1241)
1 site
T G A T C A A C T A G T
* Blocked by Dam methylation.
BclI is typically used at 50-55°C, but is 50% active at 37°C.
BstEII  (1408)
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.
PspOMI  (1434)
1 site
G G G C C C C C C G G G
ApaI  (1438)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
BssHII  (1638)
1 site
G C G C G C C G C G C G

BssHII is typically used at 50°C, but is 75% active at 37°C.
EcoRV  (1677)
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.
HpaI  (1733)
1 site
G T T A A C C A A T T G
PshAI  (2072)
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.
FspAI  (2309)
1 site
R T G C G C A Y Y A C G C G T R
PpuMI  (2334)
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.
Bpu10I  (2434)
1 site
C C T N A G C G G A N T C G