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pMCSG66 (linearized)

Linearized bacterial vector for ligation-independent cloning (LIC), with a 6xHis-2xHA-TEV leader plus a second LIC site for expressing an untagged protein.

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pMCSG66 (linearized).dna
Map and Sequence File:    Download    Open   
Sequence Author:  Midwest Center for Structural Genomics
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StyI (5178) ScaI (4426) PvuI (4316) PstI (4191) BsaI (4007) AhdI (3946) AlwNI (3469) PciI (3053) BspQI - SapI (2937) BstZ17I (2824) BsaAI (2805) PflFI - Tth111I (2798) BlpI (5201) PaeR7I - PspXI - XhoI (5279) EagI - NotI (5287) HindIII (5294) SalI (5300) Eco53kI (5309) SacI (5311) EcoRI (5313) BamHI (5319) TspMI - XmaI (5353) SmaI (5355) End (5381) Start (0) Acc65I (25) KpnI (29) BfuAI - BspMI (37) BsiWI (48) BglII (86) NdeI (122) XbaI (160) BspDI * - ClaI * (229) SgrAI (271) SphI (427) EcoNI (487) PflMI (534) BstAPI (635) MluI (952) BclI * (966) BstEII (1133) PspOMI (1159) ApaI (1163) BssHII (1363) EcoRV (1402) HpaI (1458) PshAI (1797) FspAI (2034) PpuMI (2059) Bpu10I (2159) pMCSG66 5381 bp
StyI  (5178)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
ScaI  (4426)
1 site
A G T A C T T C A T G A
PvuI  (4316)
1 site
C G A T C G G C T A G C
PstI  (4191)
1 site
C T G C A G G A C G T C
BsaI  (4007)
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  (3946)
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  (3469)
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  (3053)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspQI  (2937)
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  (2937)
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  (2824)
1 site
G T A T A C C A T A T G
BsaAI  (2805)
1 site
Y A C G T R R T G C A Y
PflFI  (2798)
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  (2798)
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.
BlpI  (5201)
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  (5279)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (5279)
1 site
V C T C G A G B B G A G C T C V
XhoI  (5279)
1 site
C T C G A G G A G C T C
EagI  (5287)
1 site
C G G C C G G C C G G C
NotI  (5287)
1 site
G C G G C C G C C G C C G G C G
HindIII  (5294)
1 site
A A G C T T T T C G A A
SalI  (5300)
1 site
G T C G A C C A G C T G
Eco53kI  (5309)
1 site
G A G C T C C T C G A G
SacI  (5311)
1 site
G A G C T C C T C G A G
EcoRI  (5313)
1 site
G A A T T C C T T A A G
BamHI  (5319)
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  (5353)
1 site
C C C G G G G G G C C C
XmaI  (5353)
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  (5355)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
End  (5381)
0 sites
Start  (0)
0 sites
Acc65I  (25)
1 site
G G T A C C C C A T G G
KpnI  (29)
1 site
G G T A C C C C A T G G
BfuAI  (37)
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  (37)
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  (48)
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  (86)
1 site
A G A T C T T C T A G A
NdeI  (122)
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  (160)
1 site
T C T A G A A G A T C T
BspDI  (229)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (229)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
SgrAI  (271)
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  (427)
1 site
G C A T G C C G T A C G
EcoNI  (487)
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  (534)
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  (635)
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  (952)
1 site
A C G C G T T G C G C A
BclI  (966)
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  (1133)
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  (1159)
1 site
G G G C C C C C C G G G
ApaI  (1163)
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  (1363)
1 site