pDEST10

Gateway® destination vector for expressing N-terminally 6xHis-tagged proteins in insect cells using the Bac-to-Bac® baculovirus system.

Sequence Author: Thermo Fisher (Invitrogen)

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BbsI (6677) PflFI - Tth111I (6148) BseRI (5881) SacII (5688) BspQI - SapI (5301) PspFI (4884) BseYI (4880) FspI (4073) PvuI (3927) BsaHI (3756) NaeI (3049) SnaBI (93) RsrII (251) ATG 6xHis TEV Site PvuII (824) MluI (1371) BssHII (1418) BbvCI (1649) TspMI - XmaI (1793) SmaI - SrfI (1795) BmgBI (1829) BfuAI - BspMI (2039) StuI (2208) SgrDI (2214) Eco53kI (2222) SacI (2224) SpeI (2227) BstBI (2243) XbaI (2248) PaeR7I - XhoI (2263) SphI (2273) Acc65I (2275) KpnI (2279) HindIII (2281) MfeI (2396) HpaI (2409) BclI * (2545) AvrII (2560) NgoMIV (3047) pDEST™10 6708 bp
BbsI  (6677)
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.
PflFI  (6148)
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  (6148)
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.
BseRI  (5881)
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.
SacII  (5688)
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.
BspQI  (5301)
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  (5301)
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.
PspFI  (4884)
1 site
C C C A G C G G G T C G
BseYI  (4880)
1 site
C C C A G C G G G T C G

After cleavage, BseYI can remain bound to DNA and alter its electrophoretic mobility.
FspI  (4073)
1 site
T G C G C A A C G C G T
PvuI  (3927)
1 site
C G A T C G G C T A G C
BsaHI  (3756)
1 site
G R C G Y C C Y G C R G

BsaHI is typically used at 37°C, but is even more active at 60°C.
NaeI  (3049)
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.
SnaBI  (93)
1 site
T A C G T A A T G C A T
RsrII  (251)
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.
PvuII  (824)
1 site
C A G C T G G T C G A C
MluI  (1371)
1 site
A C G C G T T G C G C A
BssHII  (1418)
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.
BbvCI  (1649)
1 site
C C T C A G C G G A G T C G
TspMI  (1793)
1 site
C C C G G G G G G C C C
XmaI  (1793)
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  (1795)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.