pcDNA3.2 capTEV-NT V5-DEST

Gateway® destination vector for mammalian cell expression of proteins N-terminally modified with 6xHis and V5 tags plus a cleavable biotinylation peptide.

Sequence Author: Thermo Fisher (Invitrogen)

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HomePlasmidsGateway® Cloning VectorspcDNA3.2 capTEV-NT V5-DEST
SgrDI (7919) PvuI (7368) PciI (6105) PfoI (5529) NruI (5436) RsrII (5268) PflFI - Tth111I (4870) EagI (4658) BclI * (4593) AvrII (4542) StuI (4541) CsiI - SexAI * (4309) DraIII (4019) MluI (215) SpeI (236) CMV enhancer NdeI (471) SnaBI (577) Eco53kI (803) SacI (805) AleI (875) BstEII (877) BmgBI (1945) SrfI (1979) BbvCI (2122) EcoRI (2848) BspEI (2852) EcoO109I (3737) pcDNA™3.2/capTEV-NT/V5-DEST 7921 bp
SgrDI  (7919)
1 site		 C G T C G A C G G C A G C T G C
PvuI  (7368)
1 site		 C G A T C G G C T A G C
PciI  (6105)
1 site		 A C A T G T T G T A C A
PciI is inhibited by nonionic detergents.
PfoI  (5529)
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.
NruI  (5436)
1 site		 T C G C G A A G C G C T
RsrII  (5268)
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  (4870)
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  (4870)
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.
EagI  (4658)
1 site		 C G G C C G G C C G G C
BclI  (4593)
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  (4542)
1 site		 C C T A G G G G A T C C
StuI  (4541)
1 site		 A G G C C T T C C G G A
CsiI  (4309)
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  (4309)
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.
DraIII  (4019)
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.
MluI  (215)
1 site		 A C G C G T T G C G C A
SpeI  (236)
1 site		 A C T A G T T G A T C A
NdeI  (471)
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  (577)
1 site		 T A C G T A A T G C A T
Eco53kI  (803)
1 site		 G A G C T C C T C G A G
SacI  (805)
1 site		 G A G C T C C T C G A G
AleI  (875)
1 site		 C A C N N N N G T G G T G N N N N C A C
BstEII  (877)
1 site		 G G T N A C C C C A N T G G
Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
BmgBI  (1945)
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  (1979)
1 site		 G C C C G G G C C G G G C C C G
BbvCI  (2122)
1 site		 C C T C A G C G G A G T C G
EcoRI  (2848)
1 site		 G A A T T C C T T A A G
BspEI  (2852)
1 site		 T C C G G A A G G C C T
EcoO109I  (3737)
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.
AmpR
6925 .. 7785  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   6925 .. 7716  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
6925 .. 7785  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   7717 .. 7785  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
6925 .. 7785  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
NeoR/KanR
4624 .. 5418  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
NeoR/KanR
4624 .. 5418  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from Tn5
confers resistance to neomycin, kanamycin, and G418 (Geneticin®)
CmR
2413 .. 3066  =  654 bp
218 amino acids  =  25.6 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
2413 .. 3066  =  654 bp
218 amino acids  =  25.6 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
ori
6166 .. 6754  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
6166 .. 6754  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
3786 .. 4214  =  429 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
3786 .. 4214  =  429 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
CMV enhancer
222 .. 601  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
222 .. 601  =  380 bp
human cytomegalovirus immediate early enhancer
capTEV™
860 .. 1162  =  303 bp
101 amino acids  =  10.6 kDa
7 segments
   Segment 1:  BioEase™ tag  
   860 .. 1078  =  219 bp
   73 amino acids  =  7.3 kDa
Product: composite tag for in vivo biotinylation, TEV protease cleavage, and 6xHis tagging
capTEV™
860 .. 1162  =  303 bp
101 amino acids  =  10.6 kDa
7 segments
   Segment 2:  
   1079 .. 1090  =  12 bp
   4 amino acids  =  306.3 Da
Product: composite tag for in vivo biotinylation, TEV protease cleavage, and 6xHis tagging
capTEV™
860 .. 1162  =  303 bp
101 amino acids  =  10.6 kDa
7 segments
   Segment 3:  TEV site  
   1091 .. 1111  =  21 bp
   7 amino acids  =  869.9 Da
Product: composite tag for in vivo biotinylation, TEV protease cleavage, and 6xHis tagging
capTEV™
860 .. 1162  =  303 bp
101 amino acids  =  10.6 kDa
7 segments
   Segment 4:  
   1112 .. 1117  =  6 bp
   2 amino acids  =  259.3 Da
Product: composite tag for in vivo biotinylation, TEV protease cleavage, and 6xHis tagging
capTEV™
860 .. 1162  =  303 bp
101 amino acids  =  10.6 kDa
7 segments
   Segment 5:  TEV site  
   1118 .. 1138  =  21 bp
   7 amino acids  =  869.9 Da
Product: composite tag for in vivo biotinylation, TEV protease cleavage, and 6xHis tagging
capTEV™
860 .. 1162  =  303 bp
101 amino acids  =  10.6 kDa
7 segments
   Segment 6:  
   1139 .. 1144  =  6 bp
   2 amino acids  =  259.3 Da
Product: composite tag for in vivo biotinylation, TEV protease cleavage, and 6xHis tagging
capTEV™
860 .. 1162  =  303 bp
101 amino acids  =  10.6 kDa
7 segments
   Segment 7:  6xHis  
   1145 .. 1162  =  18 bp
   6 amino acids  =  840.9 Da
Product: composite tag for in vivo biotinylation, TEV protease cleavage, and 6xHis tagging
capTEV™
860 .. 1162  =  303 bp
101 amino acids  =  10.6 kDa
7 segments
Product: composite tag for in vivo biotinylation, TEV protease cleavage, and 6xHis tagging
V5 tag
1163 .. 1204  =  42 bp
14 amino acids  =  1.4 kDa
Product: epitope tag from simian virus 5
V5 tag
1163 .. 1204  =  42 bp
14 amino acids  =  1.4 kDa
Product: epitope tag from simian virus 5
SV40 promoter
4228 .. 4557  =  330 bp
SV40 enhancer and early promoter
SV40 promoter
4228 .. 4557  =  330 bp
SV40 enhancer and early promoter
ccdB
1758 .. 2063  =  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
1758 .. 2063  =  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 promoter
602 .. 805  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
CMV promoter
602 .. 805  =  204 bp
human cytomegalovirus (CMV) immediate early promoter
attR1
1205 .. 1329  =  125 bp
recombination site for the Gateway® LR reaction
attR1
1205 .. 1329  =  125 bp
recombination site for the Gateway® LR reaction
attR2
3347 .. 3471  =  125 bp
recombination site for the Gateway® LR reaction
attR2
3347 .. 3471  =  125 bp
recombination site for the Gateway® LR reaction
SV40 poly(A) signal
5594 .. 5715  =  122 bp
SV40 polyadenylation signal
SV40 poly(A) signal
5594 .. 5715  =  122 bp
SV40 polyadenylation signal
AmpR promoter
7786 .. 7890  =  105 bp
AmpR promoter
7786 .. 7890  =  105 bp
cat promoter
3067 .. 3169  =  103 bp
promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase
cat promoter
3067 .. 3169  =  103 bp
promoter of the E. coli cat gene encoding chloramphenicol acetyltransferase
HSV TK poly(A) signal
3536 .. 3584  =  49 bp
herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)
HSV TK poly(A) signal
3536 .. 3584  =  49 bp
herpes simplex virus thymidine kinase polyadenylation signal (Cole and Stacy, 1985)
SV40 ori
4408 .. 4543  =  136 bp
SV40 origin of replication
SV40 ori
4408 .. 4543  =  136 bp
SV40 origin of replication
ORF:  4624 .. 5418  =  795 bp
ORF:  264 amino acids  =  29.0 kDa
ORF:  761 .. 1240  =  480 bp
ORF:  159 amino acids  =  17.3 kDa
ORF:  4796 .. 5182  =  387 bp
ORF:  128 amino acids  =  14.6 kDa
ORF:  7055 .. 7321  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  812 .. 1162  =  351 bp
ORF:  116 amino acids  =  12.7 kDa
ORF:  2383 .. 3066  =  684 bp
ORF:  227 amino acids  =  26.7 kDa
ORF:  4933 .. 5187  =  255 bp
ORF:  84 amino acids  =  9.6 kDa
ORF:  6925 .. 7785  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  1758 .. 2063  =  306 bp
ORF:  101 amino acids  =  11.7 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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