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Plasmid Files

pCR®8/GW/TOPO® (linearized)

Linearized vector with 3'-T overhangs and bound topoisomerase, for TOPO® TA cloning of PCR products to create a Gateway® entry vector.

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pCR8 GW TOPO (linearized) Sequence and MappCR8 GW TOPO (linearized).dna
Map and Sequence File   
Sequence Author:  Invitrogen (Life Technologies)
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 PciI (2129) DrdI (2027) BciVI (1931) AlwNI (1720) PaeR7I - PspXI - XhoI (1395) BsaAI (1356) BssHII (1323) NaeI (1250) NgoMIV (1248) BglI (1231) BtgZI (1157) PvuI (980) BfuAI - BspMI (946) BlpI (908) MslI (871) BpmI (851) BclI * (797) BstEII (759) NspI (2133) BspQI - SapI (2246) rrnB T2 terminator AclI (2499) BbsI (2572) HincII - HpaI (2636) AhdI (2684) AflII (2689) PspOMI (2698) EcoO109I (2699) ApaI - BanII (2702) AanI - AanI (2773) EcoRI (2806) End (2817) Start (1) EcoRI (7) AanI - AanI (43) EcoRV (126) M13 rev BseRI (229) XbaI (242) BspHI (599) pCR®8/GW/TOPO® 2816 bp
PciI  (2129)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
DrdI  (2027)
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.
BciVI  (1931)
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  (1720)
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.
PaeR7I  (1395)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (1395)
1 site
V C T C G A G B B G A G C T C V
XhoI  (1395)
1 site
C T C G A G G A G C T C
BsaAI  (1356)
1 site
Y A C G T R R T G C A Y
BssHII  (1323)
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.
NaeI  (1250)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NaeI
recognition sequence.
NgoMIV  (1248)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NgoMIV
recognition sequence.
BglI  (1231)
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.
BtgZI  (1157)
1 site
G C G A T G ( N ) 10 C G C T A C ( N ) 10 ( N ) 4

Sticky ends from different BtgZI sites may not be compatible.
After cleavage, BtgZI can remain bound to DNA and alter its
electrophoretic mobility.
BtgZI is typically used at 60°C, but is 75% active at 37°C.
PvuI  (980)
1 site
C G A T C G G C T A G C
BfuAI  (946)
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  (946)
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.
BlpI  (908)
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.
MslI  (871)
1 site
C A Y N N N N R T G G T R N N N N Y A C
BpmI  (851)
1 site
C T G G A G ( N ) 14 N N G A C C T C ( N ) 14

Efficient cleavage requires at least two copies of the BpmI
recognition sequence.
Sticky ends from different BpmI sites may not be compatible.
After cleavage, BpmI can remain bound to DNA and alter its
electrophoretic mobility.
BpmI quickly loses activity at 37°C.
BclI  (797)
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  (759)
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.
NspI  (2133)
1 site
R C A T G Y Y G T A C R
BspQI  (2246)
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  (2246)
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.
AclI  (2499)
1 site
A A C G T T T T G C A A
BbsI  (2572)
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.
HincII  (2636)
1 site
G T Y R A C C A R Y T G
HpaI  (2636)
1 site
G T T A A C C A A T T G
AhdI  (2684)
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  (2689)
1 site
C T T A A G G A A T T C

The sticky ends produced by AflII are hard to ligate.
PspOMI  (2698)
1 site
G G G C C C C C C G G G
EcoO109I  (2699)
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  (2702)
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  (2702)
1 site
G R G C Y C C Y C G R G

Sticky ends from different BanII sites may not be compatible.
AanI  (2773)
2 sites
T T A T A A A A T A T T
AanI  (2773)
2 sites
T T A T A A A A T A T T
EcoRI  (2806)
2 sites
G A A T T C C T T A A G
End  (2817)
0 sites
Start  (1)
0 sites
EcoRI  (7)
2 sites
G A A T T C C T T A A G
AanI  (43)
2 sites
T T A T A A A A T A T T
AanI  (43)
2 sites
T T A T A A A A T A T T
EcoRV  (126)
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  (229)
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.
XbaI  (242)
1 site
T C T A G A A G A T C T
BspHI  (599)
1 site
T C A T G A A G T A C T
SmR
601 .. 1392  =  792 bp
263 amino acids  =  29.3 kDa
Product: aminoglycoside adenylyltransferase
(Murphy, 1985)
confers resistance to spectinomycin and
streptomycin
SmR
601 .. 1392  =  792 bp
263 amino acids  =  29.3 kDa
Product: aminoglycoside adenylyltransferase
(Murphy, 1985)
confers resistance to spectinomycin and
streptomycin
ori
1485 .. 2073  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
1485 .. 2073  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
attL2
14 .. 113  =  100 bp
recombination site for the Gateway® LR reaction
attL2
14 .. 113  =  100 bp
recombination site for the Gateway® LR reaction
attL1
2704 .. 2803  =  100 bp
recombination site for the Gateway® LR reaction
attL1
2704 .. 2803  =  100 bp
recombination site for the Gateway® LR reaction
rrnB T1 terminator
2522 .. 2608  =  87 bp
transcription terminator T1 from the E. coli rrnB
gene
rrnB T1 terminator
2522 .. 2608  =  87 bp
transcription terminator T1 from the E. coli rrnB
gene
rrnB T2 terminator
2403 .. 2430  =  28 bp
transcription terminator T2 from the E. coli rrnB
gene
rrnB T2 terminator
2403 .. 2430  =  28 bp
transcription terminator T2 from the E. coli rrnB
gene
T7 promoter
131 .. 149  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
131 .. 149  =  19 bp
promoter for bacteriophage T7 RNA polymerase
M13 rev
154 .. 170  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 rev
154 .. 170  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 fwd
2672 .. 2688  =  17 bp
common sequencing primer, one of multiple similar
variants
M13 fwd
2672 .. 2688  =  17 bp
common sequencing primer, one of multiple similar
variants
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