pcDNA3.1D V5-His-TOPO

Mammalian directional TOPO® cloning vector for expressing C-terminally V5- and 6xHis-tagged proteins.

Sequence Author: Thermo Fisher (Invitrogen)

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HomePlasmidsTOPO Cloning VectorspcDNA3.1D V5-His-TOPO
SgrDI (4574) SspI (4457) ScaI (4133) PvuI (4023) AhdI (3653) PciI (2760) BstZ17I (2381) BsmI (2329) PfoI (2184) RsrII (1923) BssHII (1804) BglII (4588) MfeI (4737) Bpu10I (4756) SpeI (4825) CMV enhancer NdeI (5060) SnaBI (5166) HindIII (5478) Acc65I (5484) KpnI (5488) BamHI (5496) EcoRV (24) BstXI (34) NotI (39) PaeR7I - PspXI - XhoI (45) XbaI (51) EcoO109I - PspOMI (57) ApaI (61) SacII (64) BstBI (68) AgeI (118) 6xHis PmeI (148) BbsI (359) DraIII (673) CsiI - SexAI * (964) StuI (1196) AvrII (1197) TspMI - XmaI (1218) SmaI (1220) BsaBI * (1266) KasI (1406) NarI (1407) SfoI (1408) PluTI (1410) MscI (1489) PflFI - Tth111I (1525) pcDNA3.1D/V5-His-TOPO® 5510 bp
SgrDI  (4574)
1 site		 C G T C G A C G G C A G C T G C
SspI  (4457)
1 site		 A A T A T T T T A T A A
ScaI  (4133)
1 site		 A G T A C T T C A T G A
PvuI  (4023)
1 site		 C G A T C G G C T A G C
AhdI  (3653)
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  (2760)
1 site		 A C A T G T T G T A C A
PciI is inhibited by nonionic detergents.
BstZ17I  (2381)
1 site		 G T A T A C C A T A T G
BsmI  (2329)
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  (2184)
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.
RsrII  (1923)
1 site		 C G G W C C G G C C W G G C
Efficient cleavage requires at least two copies of the RsrII recognition sequence.
Sticky ends from different RsrII sites may not be compatible.
For full activity, add fresh DTT.
BssHII  (1804)
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.
BglII  (4588)
1 site		 A G A T C T T C T A G A
MfeI  (4737)
1 site		 C A A T T G G T T A A C
Bpu10I  (4756)
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  (4825)
1 site		 A C T A G T T G A T C A
NdeI  (5060)
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  (5166)
1 site		 T A C G T A A T G C A T
HindIII  (5478)
1 site		 A A G C T T T T C G A A
Acc65I  (5484)
1 site		 G G T A C C C C A T G G
KpnI  (5488)
1 site		 G G T A C C C C A T G G
BamHI  (5496)
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.
EcoRV  (24)
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.
BstXI  (34)
1 site		 C C A N N N N N N T G G G G T N N N N N N A C C
Sticky ends from different BstXI sites may not be compatible.
NotI  (39)
1 site		 G C G G C C G C C G C C G G C G
PaeR7I  (45)
1 site		 C T C G A G G A G C T C
PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (45)
1 site		 V C T C G A G B B G A G C T C V
XhoI  (45)
1 site		 C T C G A G G A G C T C
XbaI  (51)
1 site		 T C T A G A A G A T C T
EcoO109I  (57)
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.
PspOMI  (57)
1 site		 G G G C C C C C C G G G
ApaI  (61)
1 site		 G G G C C C C C C G G G
ApaI can be used between 25°C and 37°C.
SacII  (64)
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.
BstBI  (68)
1 site		 T T C G A A A A G C T T
AgeI  (118)
1 site		 A C C G G T T G G C C A
PmeI  (148)
1 site		 G T T T A A A C C A A A T T T G
BbsI  (359)
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.
DraIII  (673)
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.
CsiI  (964)
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  (964)
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  (1196)
1 site		 A G G C C T T C C G G A
AvrII  (1197)
1 site		 C C T A G G G G A T C C
TspMI  (1218)
1 site		 C C C G G G G G G C C C
XmaI  (1218)
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  (1220)
1 site		 C C C G G G G G G C C C
SmaI can be used at 37°C for brief incubations.
BsaBI  (1266)
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.
KasI  (1406)
1 site		 G G C G C C C C G C G G
NarI  (1407)
1 site		 G G C G C C C C G C G G
Efficient cleavage requires at least two copies of the NarI recognition sequence.
SfoI  (1408)
1 site		 G G C G C C C C G C G G
PluTI  (1410)
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.
MscI  (1489)
1 site		 T G G C C A A C C G G T
PflFI  (1525)
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  (1525)
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.
AmpR
3580 .. 4440  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   3580 .. 4371  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3580 .. 4440  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   4372 .. 4440  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
3580 .. 4440  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
NeoR/KanR
1279 .. 2073  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
NeoR/KanR
1279 .. 2073  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
ori
2821 .. 3409  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2821 .. 3409  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
440 .. 868  =  429 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
440 .. 868  =  429 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
CMV enhancer
4811 .. 5190  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
4811 .. 5190  =  380 bp
human cytomegalovirus immediate early enhancer
SV40 promoter
882 .. 1212  =  331 bp
SV40 enhancer and early promoter
SV40 promoter
882 .. 1212  =  331 bp
SV40 enhancer and early promoter
bGH poly(A) signal
170 .. 394  =  225 bp
bovine growth hormone polyadenylation signal
bGH poly(A) signal
170 .. 394  =  225 bp
bovine growth hormone polyadenylation signal
CMV promoter
5191 .. 5394  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
5191 .. 5394  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
SV40 poly(A) signal
2249 .. 2370  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
2249 .. 2370  =  122 bp
SV40 polyadenylation signal
AmpR promoter
4441 .. 4545  =  105 bp
AmpR promoter
4441 .. 4545  =  105 bp
V5 tag
73 .. 114  =  42 bp
14 amino acids  =  1.4 kDa
Product: epitope tag from simian virus 5
V5 tag
73 .. 114  =  42 bp
14 amino acids  =  1.4 kDa
Product: epitope tag from simian virus 5
6xHis
124 .. 141  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
124 .. 141  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
T7 promoter
5439 .. 5457  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
5439 .. 5457  =  19 bp
promoter for bacteriophage T7 RNA polymerase
SV40 ori
1063 .. 1198  =  136 bp
SV40 origin of replication
SV40 ori
1063 .. 1198  =  136 bp
SV40 origin of replication
ORF:  391 .. 657  =  267 bp
ORF:  88 amino acids  =  9.3 kDa
ORF:  1279 .. 2073  =  795 bp
ORF:  264 amino acids  =  29.0 kDa
ORF:  1451 .. 1837  =  387 bp
ORF:  128 amino acids  =  14.6 kDa
ORF:  3710 .. 3976  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  1588 .. 1842  =  255 bp
ORF:  84 amino acids  =  9.6 kDa
ORF:  3580 .. 4440  =  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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