pMCentr1 (linearized)

Linearized vector for ligation-independent cloning (LIC) to create a Gateway® entry vector, with a Zeocin™ resistance marker and a TEV cleavage site.

Sequence Author: Midwest Center for Structural Genomics

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PciI (1415) DrdI (1313) BssSI - BssSαI (1242) BciVI (1217) AlwNI (1006) AcuI - Eco57MI (873) BsaAI - PmlI (683) NcoI - StyI (612) MscI (611) BssHII - MauBI (574) AatII (572) ZraI (570) SmaI (524) TspMI - XmaI (522) SgrAI (496) BmgBI (480) CsiI - SexAI * (446) BanI (440) FseI (342) BglI (312) NspI (1419) BspQI - SapI (1532) BtsI - BtsαI (1635) TsoI (1678) AclI (1785) BbsI (1858) HpaI (1922) AhdI (1970) AflII (1975) PspOMI (1984) EcoO109I (1985) ApaI - BanII (1988) MslI (2039) End (2100) Start (0) EcoRV (129) BseRI (241) DraIII (264) EagI (279) pMCentr1 2100 bp
PciI  (1415)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
DrdI  (1313)
1 site
G A C N N N N N N G T C C T G N N N N N N C A G

Sticky ends from different DrdI sites may not be compatible.
BssSI  (1242)
1 site
C A C G A G G T G C T C
BssSαI  (1242)
1 site
C A C G A G G T G C T C
BciVI  (1217)
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.
AlwNI  (1006)
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.
AcuI  (873)
1 site
C T G A A G ( N ) 14 N N G A C T T C ( N ) 14

Cleavage may be enhanced when more than one copy of the AcuI recognition sequence is present.
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).
Eco57MI  (873)
1 site
C T G R A G ( N ) 14 N N G A C Y T C ( N ) 14

Sticky ends from different Eco57MI sites may not be compatible.
After cleavage, Eco57MI can remain bound to DNA and alter its electrophoretic mobility.
For full activity, add fresh S-adenosylmethionine (SAM).
BsaAI  (683)
1 site
Y A C G T R R T G C A Y
PmlI  (683)
1 site
C A C G T G G T G C A C
NcoI  (612)
1 site
C C A T G G G G T A C C
StyI  (612)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
MscI  (611)
1 site
T G G C C A A C C G G T
BssHII  (574)
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.
MauBI  (574)
1 site
C G C G C G C G G C G C G C G C
AatII  (572)
1 site
G A C G T C C T G C A G
ZraI  (570)
1 site
G A C G T C C T G C A G
SmaI  (524)
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  (522)
1 site
C C C G G G G G G C C C
XmaI  (522)
1 site
C C C G G G G G G C C C

Cleavage may be enhanced when more than one copy of the XmaI recognition sequence is present.
SgrAI  (496)
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.
BmgBI  (480)
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.
CsiI  (446)
1 site
A C C W G G T T G G W C C A

Sticky ends from different CsiI sites may not be compatible.
SexAI  (446)
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.
BanI  (440)
1 site
G G Y R C C C C R Y G G

Sticky ends from different BanI sites may not be compatible.
FseI  (342)
1 site
G G C C G G C C C C G G C C G G

FseI gradually loses activity when stored at -20°C.
BglI  (312)
1 site
G C C N N N N N G G C C G G N N N N N C C G

Sticky ends from different BglI sites may not be compatible.
NspI  (1419)
1 site
R C A T G Y Y G T A C R
BspQI  (1532)
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  (1532)
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.
BtsI  (1635)
1 site
G C A G T G N N C G T C A C
BtsαI  (1635)
1 site
G C A G T G N N C G T C A C

Sticky ends from different BtsαI sites may not be compatible.
TsoI  (1678)
1 site
T A R C C A ( N ) 9 N N A T Y G G T ( N ) 9

Sticky ends from different TsoI sites may not be compatible.
After cleavage, TsoI can remain bound to DNA and alter its electrophoretic mobility.
For full activity, add fresh S-adenosylmethionine (SAM).
AclI  (1785)
1 site
A A C G T T T T G C A A
BbsI  (1858)
1 site
G A A G A C N N C T T C T G N N ( N ) 4

Sticky ends from different BbsI sites may not be compatible.
BbsI gradually loses activity when stored at -20°C.
HpaI  (1922)
1 site
G T T A A C C A A T T G
AhdI  (1970)
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.
AflII  (1975)
1 site
C T T A A G G A A T T C
PspOMI  (1984)
1 site
G G G C C C C C C G G G
EcoO109I  (1985)
1 site
R G G N C C Y Y C C N G G R

Sticky ends from different EcoO109I sites may not be compatible.
ApaI  (1988)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
BanII  (1988)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
MslI  (2039)
1 site
C A Y N N N N R T G G T R N N N N Y A C
End  (2100)
0 sites
Start  (0)
0 sites
EcoRV  (129)
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.
BseRI  (241)
1 site
G A G G A G ( N ) 8 N N C T C C T C ( N ) 8

Sticky ends from different BseRI sites may not be compatible.
BseRI quickly loses activity at 37°C.
Prolonged incubation with BseRI may lead to degradation of the DNA.
DraIII  (264)
1 site
C A C N N N G T G G T G N N N C A C

Sticky ends from different DraIII sites may not be compatible.
EagI  (279)
1 site
C G G C C G G C C G G C
ori
771 .. 1359  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
771 .. 1359  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
BleoR
241 .. 615  =  375 bp
124 amino acids  =  13.8 kDa
Product: antibiotic-binding protein
confers resistance to bleomycin, phleomycin, and Zeocin™
BleoR
241 .. 615  =  375 bp
124 amino acids  =  13.8 kDa
Product: antibiotic-binding protein
confers resistance to bleomycin, phleomycin, and Zeocin™
attL2
17 .. 116  =  100 bp
recombination site for the Gateway® LR reaction
attL2
17 .. 116  =  100 bp
recombination site for the Gateway® LR reaction
attL1
1990 .. 2089  =  100 bp
recombination site for the Gateway® LR reaction
attL1
1990 .. 2089  =  100 bp
recombination site for the Gateway® LR reaction
rrnB T1 terminator
1808 .. 1894  =  87 bp
transcription terminator T1 from the E. coli rrnB gene
rrnB T1 terminator
1808 .. 1894  =  87 bp
transcription terminator T1 from the E. coli rrnB gene
EM7 promoter
634 .. 681  =  48 bp
synthetic bacterial promoter
EM7 promoter
634 .. 681  =  48 bp
synthetic bacterial promoter
rrnB T2 terminator
1689 .. 1716  =  28 bp
transcription terminator T2 from the E. coli rrnB gene
rrnB T2 terminator
1689 .. 1716  =  28 bp
transcription terminator T2 from the E. coli rrnB gene
TEV site
2092 .. 2112  =  21 bp
7 amino acids  =  900.0 Da
Product: tobacco etch virus (TEV) protease recognition and cleavage site
TEV site
2092 .. 2112  =  21 bp
7 amino acids  =  900.0 Da
Product: tobacco etch virus (TEV) protease recognition and cleavage site
T7 promoter
134 .. 152  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
134 .. 152  =  19 bp
promoter for bacteriophage T7 RNA polymerase
M13 rev
157 .. 173  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
157 .. 173  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
1958 .. 1974  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
1958 .. 1974  =  17 bp
common sequencing primer, one of multiple similar variants
ORF:  335 .. 622  =  288 bp
ORF:  95 amino acids  =  10.1 kDa
ORF:  237 .. 467  =  231 bp
ORF:  76 amino acids  =  8.2 kDa
ORF:  241 .. 615  =  375 bp
ORF:  124 amino acids  =  13.8 kDa
ORF:  1815 .. 2114  =  300 bp
ORF:  99 amino acids  =  11.3 kDa  (no start codon)
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