pDEST27

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

Sequence Author: Thermo Fisher (Invitrogen)

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HomePlasmidsGateway® Cloning VectorspDEST27
AhdI (6846) PfoI (5807) RsrII (5386) PluTI (4873) SfoI (4871) NarI (4870) KasI (4869) AflII (4724) HindIII (4692) StuI (4675) BseRI (4672) SfiI (4629) CsiI - SexAI * (4443) KpnI (4330) Acc65I (4326) SnaBI (294) Eco53kI (520) SacI (522) BbsI (588) SacII (626) EcoNI (642) BsgI (918) SwaI (1059) XbaI * (1304) NotI (1439) EcoRI (1759) PasI - PflMI * (1991) BglII (2206) BstZ17I (2294) BbvCI (2484) AvaI - BsoBI - TspMI - XmaI (2628) SmaI - SrfI (2630) BmgBI (2664) SalI (2887) M13 fwd NheI (3117) BmtI (3121) MfeI (3486) HpaI (3499) DraIII (3993) pDEST™27 8123 bp
AhdI  (6846)
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.
PfoI  (5807)
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  (5386)
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.
PluTI  (4873)
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  (4871)
1 site		 G G C G C C C C G C G G
NarI  (4870)
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  (4869)
1 site		 G G C G C C C C G C G G
AflII  (4724)
1 site		 C T T A A G G A A T T C
HindIII  (4692)
1 site		 A A G C T T T T C G A A
StuI  (4675)
1 site		 A G G C C T T C C G G A
BseRI  (4672)
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.
SfiI  (4629)
1 site		 G G C C N N N N N G G C C C C G G N N N N N C C G G
Efficient cleavage requires at least two copies of the SfiI recognition sequence.
Sticky ends from different SfiI sites may not be compatible.
CsiI  (4443)
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  (4443)
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.
KpnI  (4330)
1 site		 G G T A C C C C A T G G
Acc65I  (4326)
1 site		 G G T A C C C C A T G G
SnaBI  (294)
1 site		 T A C G T A A T G C A T
Eco53kI  (520)
1 site		 G A G C T C C T C G A G
SacI  (522)
1 site		 G A G C T C C T C G A G
BbsI  (588)
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.
SacII  (626)
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.
EcoNI  (642)
1 site		 C C T N N N N N A G G G G A N N N N N T C C
The 1-base overhangs produced by EcoNI may be hard to ligate.
Sticky ends from different EcoNI sites may not be compatible.
BsgI  (918)
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).
SwaI  (1059)
1 site		 A T T T A A A T T A A A T T T A
SwaI is typically used at 25°C, but is 50% active at 37°C.
XbaI  (1304)
1 site		 T C T A G A A G A T C T
* Blocked by Dam methylation.
NotI  (1439)
1 site		 G C G G C C G C C G C C G G C G
EcoRI  (1759)
1 site		 G A A T T C C T T A A G
PasI  (1991)
1 site		 C C C W G G G G G G W C C C
Sticky ends from different PasI sites may not be compatible.
PflMI  (1991)
1 site		 C C A N N N N N T G G G G T N N N N N A C C
* Blocked by Dcm methylation.
Sticky ends from different PflMI sites may not be compatible.
BglII  (2206)
1 site		 A G A T C T T C T A G A
BstZ17I  (2294)
1 site		 G T A T A C C A T A T G
BbvCI  (2484)
1 site		 C C T C A G C G G A G T C G
AvaI  (2628)
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  (2628)
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  (2628)
1 site		 C C C G G G G G G C C C
XmaI  (2628)
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  (2630)
1 site		 C C C G G G G G G C C C
SmaI can be used at 37°C for brief incubations.
SrfI  (2630)
1 site		 G C C C G G G C C G G G C C C G
BmgBI  (2664)
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.
SalI  (2887)
1 site		 G T C G A C C A G C T G
NheI  (3117)
1 site		 G C T A G C C G A T C G
BmtI  (3121)
1 site		 G C T A G C C G A T C G
MfeI  (3486)
1 site		 C A A T T G G T T A A C
HpaI  (3499)
1 site		 G T T A A C C A A T T G
DraIII  (3993)
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.
AmpR
6059 .. 6919  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   6059 .. 6127  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
6059 .. 6919  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   6128 .. 6919  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
6059 .. 6919  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
NeoR/KanR
4742 .. 5536  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
NeoR/KanR
4742 .. 5536  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
CmR
1546 .. 2205  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
1546 .. 2205  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
GST
632 .. 1285  =  654 bp
218 amino acids  =  25.5 kDa
Product: glutathione S-transferase from Schistosoma japonicum
GST
632 .. 1285  =  654 bp
218 amino acids  =  25.5 kDa
Product: glutathione S-transferase from Schistosoma japonicum
ori
7090 .. 7678  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
7090 .. 7678  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
3760 .. 4215  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
3760 .. 4215  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
SV40 promoter
4334 .. 4691  =  358 bp
SV40 enhancer and early promoter
SV40 promoter
4334 .. 4691  =  358 bp
SV40 enhancer and early promoter
ccdB
2547 .. 2852  =  306 bp
101 amino acids  =  11.7 kDa
Product: CcdB, a bacterial toxin that poisons DNA gyrase
Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains.
ccdB
2547 .. 2852  =  306 bp
101 amino acids  =  11.7 kDa
Product: CcdB, a bacterial toxin that poisons DNA gyrase
Plasmids containing the ccdB gene cannot be propagated in standard E. coli strains.
CMV enhancer
15 .. 318  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
15 .. 318  =  304 bp
human cytomegalovirus immediate early enhancer
CMV promoter
319 .. 522  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
319 .. 522  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
SV40 poly(A) signal
3500 .. 3634  =  135 bp
SV40 polyadenylation signal
SV40 poly(A) signal
3500 .. 3634  =  135 bp
SV40 polyadenylation signal
attR1
1313 .. 1437  =  125 bp
recombination site for the Gateway® LR reaction
attR1
1313 .. 1437  =  125 bp
recombination site for the Gateway® LR reaction
attR2
2893 .. 3017  =  125 bp
recombination site for the Gateway® LR reaction
attR2
2893 .. 3017  =  125 bp
recombination site for the Gateway® LR reaction
AmpR promoter
5954 .. 6058  =  105 bp
AmpR promoter
5954 .. 6058  =  105 bp
poly(A) signal
5600 .. 5648  =  49 bp
synthetic polyadenylation signal
poly(A) signal
5600 .. 5648  =  49 bp
synthetic polyadenylation signal
lac UV5 promoter
1462 .. 1492  =  31 bp
3 segments
   Segment 1:  -35  
   1462 .. 1467  =  6 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
1462 .. 1492  =  31 bp
3 segments
   Segment 2:  
   1468 .. 1485  =  18 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
1462 .. 1492  =  31 bp
3 segments
   Segment 3:  -10  
   1486 .. 1492  =  7 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
1462 .. 1492  =  31 bp
3 segments
E. coli lac promoter with an "up" mutation
T7 promoter
3050 .. 3068  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
3050 .. 3068  =  19 bp
promoter for bacteriophage T7 RNA polymerase
M13 fwd
3079 .. 3095  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
3079 .. 3095  =  17 bp
common sequencing primer, one of multiple similar variants
SV40 ori
4542 .. 4677  =  136 bp
SV40 origin of replication
SV40 ori
4542 .. 4677  =  136 bp
SV40 origin of replication
ORF:  1546 .. 2205  =  660 bp
ORF:  219 amino acids  =  25.7 kDa
ORF:  632 .. 1348  =  717 bp
ORF:  238 amino acids  =  27.8 kDa
ORF:  4742 .. 5536  =  795 bp
ORF:  264 amino acids  =  29.0 kDa
ORF:  6059 .. 6919  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  8078 .. 212  =  258 bp
ORF:  85 amino acids  =  9.6 kDa
ORF:  2547 .. 2852  =  306 bp
ORF:  101 amino acids  =  11.7 kDa
ORF:  4914 .. 5300  =  387 bp
ORF:  128 amino acids  =  14.6 kDa
ORF:  5051 .. 5587  =  537 bp
ORF:  178 amino acids  =  19.8 kDa
ORF:  6523 .. 6789  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
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Individual Sequences & Maps

pAd BLOCK-iT-DEST
pDEST R4-R3 Vector II
pENTR11
pHELLSGATE 4
pAd CMV V5-DEST
pDEST10
pENTR1A
pHELLSGATE 8
pAd PL-DEST
pDEST14
pENTR2B
pIB V5-His-DEST
pBAD-DEST49
pDEST15
pENTR3C
pLenti4 BLOCK-iT-DEST
pcDNA-DEST40
pDEST17
pENTR4
pLenti4 V5-DEST
pcDNA-DEST47
pDEST20
pET-53-DEST
pLenti6.2 C-Lumio V5-DEST
pcDNA-DEST53
pDEST24
pET-54-DEST
pLenti6.2 N-Lumio V5-DEST
pcDNA3.1 nV5-DEST
pDEST26
pET-55-DEST
pLenti6.2 V5-DEST
pcDNA3.2 capTEV-CT V5-DEST
pDEST27
pET-56-DEST
pLenti6.3 TO V5-DEST
pcDNA3.2 capTEV-NT V5-DEST
pDEST8
pET-57-DEST
pLenti6.3 V5-DEST
pcDNA3.2 V5-DEST
pDONR221 P5-P2
pET-58-DEST
pLenti6.4 R4R2 V5-DEST
pcDNA6.2 C-EmGFP-DEST
pDONR P2r-P3
pET-59-DEST
pLenti6 BLOCK-iT-DEST
pcDNA6.2 C-YFP-DEST
pDONR P4-P1r
pET-60-DEST
pLenti6 capTEV-CT-DEST
pcDNA6.2 cGeneBLAzer-DEST
pDONR Zeo
pET-61-DEST
pLenti6 capTEV-NT-DEST1
pcDNA6.2 cLumio-DEST
pDONR201
pET-62-DEST
pLenti6 UbC V5-DEST
pcDNA6.2 EmGFP-Bsd V5-DEST
pDONR207
pET-DEST42
pLenti6 V5-DEST
pcDNA6.2 N-EmGFP-DEST
pDONR221
pET104-DEST
pLenti7.3 V5-DEST
pcDNA6.2 N-YFP-DEST
pDONR221 P1-P4
pET104.1-DEST
pMT-DEST48
pcDNA6.2 nGeneBLAzer-DEST
pDONR221 P1-P5r
pET160-DEST
pMT BioEase-DEST
pcDNA6.2 nLumio-DEST
pDONR221 P3-P2
pET161-DEST
pSCREEN-iT lacZ-DEST
pcDNA6.2 V5-DEST
pDONR221 P4r-P3r
pEXP1-DEST
pT-REx-DEST30
pcDNA6.2 V5-pL-DEST
pDONR221 P5-P4
pEXP2-DEST
pT-REx-DEST31
pCMV SPORT6
pDONR222
pEXP3-DEST
pYES-DEST52
pCMV SPORT6.1
pDONR223
pEXP4-DEST
Reading Frame Cassette A
pCOLA-2-DEST
pEF-DEST51
pHELLSGATE
Reading Frame Cassette B
pCOLA-3-DEST
pEF5 FRT V5-DEST
pHELLSGATE 12
Reading Frame Cassette C.1
pDEST R4-R3
pENTR GeneBLAzer

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