pcDNA3.2 V5-DEST

Gateway® destination vector for high-level expression of C-terminally V5-tagged proteins in mammalian cells.

Sequence Author: Thermo Fisher (Invitrogen)

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SgrDI (7709) PvuI (7158) PciI (5895) PfoI (5319) RsrII (5058) PflFI - Tth111I (4660) PluTI (4545) SfoI (4543) NarI (4542) KasI (4541) EagI (4448) BclI * (4383) AvrII (4332) StuI (4331) CsiI - SexAI * (4099) MfeI (161) SpeI (249) CMV enhancer NdeI (484) SnaBI (590) Eco53kI (816) SacI (818) BstXI (1572) BmgBI (1651) SrfI (1685) BbvCI (1828) BamHI (2099) EcoRI (2552) BspEI (2556) XbaI * (3179) PspOMI (3185) ApaI (3189) SacII (3192) BstBI (3196) AgeI (3246) PmeI (3264) DraIII (3809) pcDNA™3.2/V5-DEST 7711 bp
SgrDI  (7709)
1 site
C G T C G A C G G C A G C T G C
PvuI  (7158)
1 site
C G A T C G G C T A G C
PciI  (5895)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
PfoI  (5319)
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  (5058)
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.
PflFI  (4660)
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  (4660)
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.
PluTI  (4545)
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  (4543)
1 site
G G C G C C C C G C G G
NarI  (4542)
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.
KasI  (4541)
1 site
G G C G C C C C G C G G
EagI  (4448)
1 site
C G G C C G G C C G G C
BclI  (4383)
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.
AvrII  (4332)
1 site
C C T A G G G G A T C C
StuI  (4331)
1 site
A G G C C T T C C G G A
CsiI  (4099)
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  (4099)
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.
MfeI  (161)
1 site
C A A T T G G T T A A C
SpeI  (249)
1 site
A C T A G T T G A T C A
NdeI  (484)
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  (590)
1 site
T A C G T A A T G C A T
Eco53kI  (816)
1 site
G A G C T C C T C G A G
SacI  (818)
1 site
G A G C T C C T C G A G
BstXI  (1572)
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.
BmgBI  (1651)
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.
SrfI  (1685)
1 site
G C C C G G G C C G G G C C C G
BbvCI  (1828)
1 site
C C T C A G C G G A G T C G
BamHI  (2099)
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.
EcoRI  (2552)
1 site
G A A T T C C T T A A G
BspEI  (2556)
1 site
T C C G G A A G G C C T
XbaI  (3179)
1 site
T C T A G A A G A