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pMCSG12 (linearized)

Linearized bacterial coexpression vector for ligation-independent cloning (LIC), with a 6xHis-S-loop-TEV leader and a p15A origin. 

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pMCSG12 (linearized).dna
Map and Sequence File:    Download    Open   
Sequence Author:  Midwest Center for Structural Genomics
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EcoNI (3684) BstAPI (3540) ApaLI (3236) AflIII - MluI (3216) BclI * (3202) BstEII (3034) ApaI (3013) PspOMI (3009) BciVI (2764) HpaI (2714) PluTI (2581) SfoI (2579) NarI * (2578) KasI (2577) AcuI (2356) XbaI (2296) NspI (2033) BssS α I (1831) SacII (1707) SgrAI (1541) NcoI (3780) BfuAI - BspMI (3835) AscI (3836) PstI - SbfI (3846) AflII (3874) BsrGI (3901) NdeI (4009) BglII (4043) Acc65I (4104) KpnI (4108) End (4118) Start (0) AvaI - BsoBI - PaeR7I - PspXI - XhoI (57) BmeT110I (58) PacI (132) AvrII (136) BlpI (154) EcoO109I (181) Bsu36I (220) DrdI - PflFI - Tth111I (329) ScaI (460) MscI (610) BspEI (877) Bpu10I (1101) BsaAI (1184) AfeI (1454) NheI (1455) BmtI (1459) BstZ17I (1468) XmnI (1511) pMCSG12 4118 bp
EcoNI  (3684)
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.
BstAPI  (3540)
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.
ApaLI  (3236)
1 site
G T G C A C C A C G T G
AflIII  (3216)
1 site
A C R Y G T T G Y R C A

Sticky ends from different AflIII sites may not be compatible.
MluI  (3216)
1 site
A C G C G T T G C G C A
BclI  (3202)
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  (3034)
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.
ApaI  (3013)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PspOMI  (3009)
1 site
G G G C C C C C C G G G
BciVI  (2764)
1 site
G T A T C C ( N ) 5 N C A T A G G ( N ) 5

The 1-base overhangs produced by BciVI may be hard to ligate.
Sticky ends from different BciVI sites may not be compatible.
HpaI  (2714)
1 site
G T T A A C C A A T T G
PluTI  (2581)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the PluTI recognition sequence.
SfoI  (2579)
1 site
G G C G C C C C G C G G
NarI  (2578)
1 site
G G C G C C C C G C G G
* Blocked by Dcm methylation.
Efficient cleavage requires at least two copies of the NarI recognition sequence.
KasI  (2577)
1 site
G G C G C C C C G C G G
AcuI  (2356)
1 site
C T G A A G ( N ) 14 N N G A C T T C ( N ) 14

Efficient cleavage requires at least two copies of the AcuI recognition sequence.
Sticky ends from different AcuI sites may not be compatible.
After cleavage, AcuI can remain bound to DNA and alter its electrophoretic mobility.
For full activity, add fresh S-adenosylmethionine (SAM).
XbaI  (2296)
1 site
T C T A G A A G A T C T
NspI  (2033)
1 site
R C A T G Y Y G T A C R
BssSαI  (1831)
1 site
C A C G A G G T G C T C
SacII  (1707)
1 site
C C G C G G G G C G C C

Efficient cleavage requires at least two copies of the SacII recognition sequence.
SgrAI  (1541)
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.
NcoI  (3780)
1 site
C C A T G G G G T A C C
BfuAI  (3835)
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  (3835)
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.
AscI  (3836)
1 site
G G C G C G C C C C G C G C G G
PstI  (3846)
1 site
C T G C A G G A C G T C
SbfI  (3846)
1 site
C C T G C A G G G G A C G T C C
AflII  (3874)
1 site
C T T A A G G A A T T C
BsrGI  (3901)
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.
NdeI  (4009)
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.
BglII  (4043)
1 site
A G A T C T T C T A G A
Acc65I  (4104)
1 site
G G T