pcDNA6.2 V5 GW D-TOPO

Mammalian directional TOPO® cloning vector with a blasticidin resistance gene and a C-terminal V5 tag, for creating a Gateway® expression clone.

Sequence Author: Thermo Fisher (Invitrogen)

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HomePlasmidsTOPO Cloning VectorspcDNA6.2 V5 GW D-TOPO
SgrDI (4205) SspI (4088) ScaI (3764) AhdI (3284) PciI (2394) BspQI - SapI (2278) BstZ17I (2015) BsmI (1963) PfoI (1818) BlpI (1722) BsaBI * (1625) BsgI (1514) MfeI (4368) Bpu10I (4387) SpeI (4456) NdeI (4691) SnaBI (4797) Eco53kI (5023) SacI (5025) NotI (5146) AscI - BssHII (9) attB2 XbaI * (45) PspOMI (51) ApaI (55) BstBI (62) AgeI (112) PmeI (130) CsiI - SexAI * (965) StuI (1197) AvrII (1198) AvaI - BsoBI - TspMI - XmaI (1219) SmaI (1221) BclI * (1249) MscI (1330) BbsI (1412) PpuMI (1486) pcDNA6.2/V5/GW/D-TOPO® 5158 bp
SgrDI  (4205)
1 site		 C G T C G A C G G C A G C T G C
SspI  (4088)
1 site		 A A T A T T T T A T A A
ScaI  (3764)
1 site		 A G T A C T T C A T G A
AhdI  (3284)
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.
PciI  (2394)
1 site		 A C A T G T T G T A C A
PciI is inhibited by nonionic detergents.
BspQI  (2278)
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  (2278)
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  (2015)
1 site		 G T A T A C C A T A T G
BsmI  (1963)
1 site		 G A A T G C N C T T A C G N
Sticky ends from different BsmI sites may not be compatible.
PfoI  (1818)
1 site		 T C C N G G A A G G N C C T
Sticky ends from different PfoI sites may not be compatible.
BlpI  (1722)
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.
BsaBI  (1625)
1 site		 G A T N N N N A T C C T A N N N N T A G
* Blocked by Dam methylation.
BsgI  (1514)
1 site		 G T G C A G ( N ) 14 N N C A C G T C ( N ) 14
Efficient cleavage requires at least two copies of the BsgI recognition sequence.
Sticky ends from different BsgI sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
MfeI  (4368)
1 site		 C A A T T G G T T A A C
Bpu10I  (4387)
1 site		 C C T N A G C G G A N T C G
Cleavage may be enhanced when more than one copy of the Bpu10I recognition sequence is present.
This recognition sequence is asymmetric, so ligating sticky ends generated by Bpu10I will not always regenerate a Bpu10I site.
Sticky ends from different Bpu10I sites may not be compatible.
SpeI  (4456)
1 site		 A C T A G T T G A T C A
NdeI  (4691)
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.
SnaBI  (4797)
1 site		 T A C G T A A T G C A T
Eco53kI  (5023)
1 site		 G A G C T C C T C G A G
SacI  (5025)
1 site		 G A G C T C C T C G A G
NotI  (5146)
1 site		 G C G G C C G C C G C C G G C G
AscI  (9)
1 site		 G G C G C G C C C C G C G C G G
BssHII  (9)
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.
XbaI  (45)
1 site		 T C T A G A A G A T C T
* Blocked by Dam methylation.
PspOMI  (51)
1 site		 G G G C C C C C C G G G
ApaI  (55)
1 site		 G G G C C C C C C G G G
ApaI can be used between 25°C and 37°C.
BstBI  (62)
1 site		 T T C G A A A A G C T T
AgeI  (112)
1 site		 A C C G G T T G G C C A
PmeI  (130)
1 site		 G T T T A A A C C A A A T T T G
CsiI  (965)
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  (965)
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.
StuI  (1197)
1 site		 A G G C C T T C C G G A
AvrII  (1198)
1 site		 C C T A G G G G A T C C
AvaI  (1219)
1 site		 C Y C G R G G R G C Y C
Sticky ends from different AvaI sites may not be compatible.
BsoBI  (1219)
1 site		 C Y C G R G G R G C Y C
Sticky ends from different BsoBI sites may not be compatible.
BsoBI is typically used at 37°C, but can be used at temperatures up to 65°C.
TspMI  (1219)
1 site		 C C C G G G G G G C C C
XmaI  (1219)
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.
SmaI  (1221)
1 site		 C C C G G G G G G C C C
SmaI can be used at 37°C for brief incubations.
BclI  (1249)
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.
MscI  (1330)
1 site		 T G G C C A A C C G G T
BbsI  (1412)
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.
PpuMI  (1486)
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.
AmpR
3211 .. 4071  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   3211 .. 4002  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3211 .. 4071  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   4003 .. 4071  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3211 .. 4071  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
2455 .. 3040  =  586 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2455 .. 3040  =  586 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
442 .. 870  =  429 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
442 .. 870  =  429 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
BSD
1327 .. 1725  =  399 bp
132 amino acids  =  13.7 kDa
Product: blasticidin S deaminase
confers resistance to blasticidin
BSD
1327 .. 1725  =  399 bp
132 amino acids  =  13.7 kDa
Product: blasticidin S deaminase
confers resistance to blasticidin
CMV enhancer
4442 .. 4821  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
4442 .. 4821  =  380 bp
human cytomegalovirus immediate early enhancer
SV40 promoter
884 .. 1213  =  330 bp
SV40 enhancer and early promoter
SV40 promoter
884 .. 1213  =  330 bp
SV40 enhancer and early promoter
CMV promoter
4822 .. 5025  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
4822 .. 5025  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
SV40 poly(A) signal
1883 .. 2004  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1883 .. 2004  =  122 bp
SV40 polyadenylation signal
AmpR promoter
4072 .. 4176  =  105 bp
AmpR promoter
4072 .. 4176  =  105 bp
HSV TK poly(A) signal
192 .. 240  =  49 bp
herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)
HSV TK poly(A) signal
192 .. 240  =  49 bp
herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)
EM7 promoter
1261 .. 1308  =  48 bp
synthetic bacterial promoter
EM7 promoter
1261 .. 1308  =  48 bp
synthetic bacterial promoter
V5 tag
67 .. 108  =  42 bp
14 amino acids  =  1.4 kDa
Product: epitope tag from simian virus 5
V5 tag
67 .. 108  =  42 bp
14 amino acids  =  1.4 kDa
Product: epitope tag from simian virus 5
attB2
17 .. 41  =  25 bp
recombination site for the Gateway® BP reaction
attB2
17 .. 41  =  25 bp
recombination site for the Gateway® BP reaction
attB1
5118 .. 5142  =  25 bp
recombination site for the Gateway® BP reaction
attB1
5118 .. 5142  =  25 bp
recombination site for the Gateway® BP reaction
T7 promoter
5070 .. 5088  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
5070 .. 5088  =  19 bp
promoter for bacteriophage T7 RNA polymerase
SV40 ori
1064 .. 1199  =  136 bp
SV40 origin of replication
SV40 ori
1064 .. 1199  =  136 bp
SV40 origin of replication
ORF:  1327 .. 1725  =  399 bp
ORF:  132 amino acids  =  13.7 kDa
ORF:  3341 .. 3607  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  1273 .. 1629  =  357 bp
ORF:  118 amino acids  =  12.8 kDa
ORF:  3211 .. 4071  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
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Individual Sequences & Maps

pBAD-TOPO (linearized)
pcDNA5 FRT TO-TOPO (linearized)
pEF6 V5-His-TOPO (linearized)
pGlow-TOPO (linearized)
pBAD Thio-TOPO (linearized)
pcDNA5 FRT V5-His-TOPO (linearized)
pENTR D-TOPO
pIB V5-His-TOPO (linearized)
pBAD202 D-TOPO
pcDNA6.2 C-EmGFP-GW TOPO (linearized)
pENTR SD D-TOPO
pLenti6.3 V5-TOPO (linearized)
pBlue-TOPO (linearized)
pcDNA6.2 C-YFP-GW TOPO (linearized)
pENTR TEV D-TOPO
pLenti6 V5-D-TOPO
pcDNA3.1D V5-His-TOPO
pcDNA6.2 N-EmGFP-GW TOPO (linearized)
pENTR5'-TOPO (linearized)
pLenti7.3 V5-TOPO (linearized)
pcDNA3.2 V5 GW D-TOPO
pCR-XL-2-TOPO (linearized)
pET100 D-TOPO
pLPS-B (linearized)
pcDNA6.2 cGeneBLAzer-GW D-TOPO
pCR-XL-TOPO (linearized)
pET101 D-TOPO
pMT V5-His-TOPO (linearized)
pcDNA6.2 V5 GW D-TOPO
pCR2.1-TOPO (linearized)
pET102 D-TOPO
pOptiVEC-TOPO (linearized)
pcDNA3.1 CT-GFP-TOPO (linearized)
pCR4-TOPO (linearized)
pET151 D-TOPO
pSecTag FRT V5-His-TOPO (linearized)
pcDNA3.1 NT-GFP-TOPO (linearized)
pCR4Blunt-TOPO (linearized)
pET160 GW D-TOPO
pTOP Blunt V2 (linearized)
pcDNA3.1 V5-His-TOPO (linearized)
pCR8 GW TOPO (linearized)
pET161 GW D-TOPO
pTrcHis-TOPO (linearized)
pcDNA3.3-TOPO (linearized)
pCR-Blunt II-TOPO (linearized)
pET200 D-TOPO
pTrcHis2-TOPO (linearized)
pcDNA3.4-TOPO (linearized)
pCRII-TOPO (linearized)
pEXP5-CT TOPO (linearized)
pYES2.1 V5-His-TOPO (linearized)
pcDNA4 HisMax-TOPO (linearized)
pEF5 FRT V5-D-TOPO
pEXP5-NT TOPO (linearized)

Plasmid Sets

Basic Cloning Vectors
I.M.A.G.E. Consortium Plasmids
pET & Duet Vectors (Novagen)
TA and GC Cloning Vectors
Coronavirus Resources
Insect Cell Vectors
pGEX Vectors (GE Healthcare)
TOPO Cloning Vectors
CRISPR Plasmids
Luciferase Vectors
Plant Vectors
Viral Expression & Packaging Vectors
Fluorescent Protein Genes & Plasmids
Lucigen Vectors
Qiagen Vectors
Yeast Plasmids
Gateway® Cloning Vectors
Mammalian Expression Vectors
Structural Genomics Vectors

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