pET104-DEST

Gateway® destination vector for high-level inducible expression of biotinylated proteins in bacteria.

Sequence Author: Thermo Fisher (Invitrogen)

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HomePlasmidsGateway® Cloning VectorspET104-DEST
EcoNI (7553) BclI * (7071) ApaI (6882) PspOMI (6878) HpaI (6583) PshAI (6244) NruI (5986) FspAI (5504) PflFI - Tth111I (4739) BsaAI (4733) BspQI - SapI (4598) PciI (4481) SphI (0) BglII (189) T7 promoter XbaI (255) RBS NdeI (295) ATG AleI (312) Xpress™ tag attR1 NotI (687) lac UV5 promoter EcoRI (1007) PasI (1239) NcoI (1308) AgeI (1507) BbvCI (1711) TspMI - XmaI (1855) SmaI - SrfI (1857) BmgBI (1891) SalI (2114) PacI (2254) BlpI (2319) T7 terminator BspDI - ClaI (2567) ZraI (2672) AatII (2674) PvuI (3224) AhdI (3593) pET104-DEST 7618 bp
EcoNI  (7553)
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.
BclI  (7071)
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.
ApaI  (6882)
1 site		 G G G C C C C C C G G G
ApaI can be used between 25°C and 37°C.
PspOMI  (6878)
1 site		 G G G C C C C C C G G G
HpaI  (6583)
1 site		 G T T A A C C A A T T G
PshAI  (6244)
1 site		 G A C N N N N G T C C T G N N N N C A G
PshAI quickly loses activity at 37°C, but can be used at 25°C for long incubations.
NruI  (5986)
1 site		 T C G C G A A G C G C T
FspAI  (5504)
1 site		 R T G C G C A Y Y A C G C G T R
PflFI  (4739)
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  (4739)
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.
BsaAI  (4733)
1 site		 Y A C G T R R T G C A Y
BspQI  (4598)
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  (4598)
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.
PciI  (4481)
1 site		 A C A T G T T G T A C A
PciI is inhibited by nonionic detergents.
SphI  (0)
1 site		 G C A T G C C G T A C G
BglII  (189)
1 site		 A G A T C T T C T A G A
XbaI  (255)
1 site		 T C T A G A A G A T C T
NdeI  (295)
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.
AleI  (312)
1 site		 C A C N N N N G T G G T G N N N N C A C
NotI  (687)
1 site		 G C G G C C G C C G C C G G C G
EcoRI  (1007)
1 site		 G A A T T C C T T A A G
PasI  (1239)
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.
NcoI  (1308)
1 site		 C C A T G G G G T A C C
AgeI  (1507)
1 site		 A C C G G T T G G C C A
BbvCI  (1711)
1 site		 C C T C A G C G G A G T C G
TspMI  (1855)
1 site		 C C C G G G G G G C C C
XmaI  (1855)
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  (1857)
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  (1857)
1 site		 G C C C G G G C C G G G C C C G
BmgBI  (1891)
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  (2114)
1 site		 G T C G A C C A G C T G
PacI  (2254)
1 site		 T T A A T T A A A A T T A A T T
BlpI  (2319)
1 site		 G C T N A G C C G A N T C G
Sticky ends from different BlpI sites may not be compatible.
BspDI  (2567)
1 site		 A T C G A T T A G C T A
ClaI  (2567)
1 site		 A T C G A T T A G C T A
ZraI  (2672)
1 site		 G A C G T C C T G C A G
AatII  (2674)
1 site		 G A C G T C C T G C A G
PvuI  (3224)
1 site		 C G A T C G G C T A G C
AhdI  (3593)
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.
lacI
6358 .. 7440  =  1083 bp
360 amino acids  =  38.6 kDa
Product: lac repressor
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
lacI
6358 .. 7440  =  1083 bp
360 amino acids  =  38.6 kDa
Product: lac repressor
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
AmpR
2806 .. 3666  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
   Segment 1:  signal sequence  
   2806 .. 2874  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2806 .. 3666  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
   Segment 2:  
   2875 .. 3666  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2806 .. 3666  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
CmR
794 .. 1453  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
794 .. 1453  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
ori
3837 .. 4425  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
3837 .. 4425  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ccdB
1774 .. 2079  =  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
1774 .. 2079  =  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.
ATG
297 .. 299  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
297 .. 299  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
BioEase™ tag
300 .. 515  =  216 bp
72 amino acids  =  7.1 kDa
3 segments
   Segment 1:  
   300 .. 410  =  111 bp
   37 amino acids  =  3.6 kDa
Product: biotinylated sequence derived from the C-terminus of the α-subunit of Klebsiella pneumoniae oxaloacetate decarboxylase
contains a unique biotinylation site
BioEase™ tag
300 .. 515  =  216 bp
72 amino acids  =  7.1 kDa
3 segments
   Segment 2:  site  
   411 .. 413  =  3 bp
   1 amino acid  =  146.2 Da
Product: biotinylated sequence derived from the C-terminus of the α-subunit of Klebsiella pneumoniae oxaloacetate decarboxylase
contains a unique biotinylation site
BioEase™ tag
300 .. 515  =  216 bp
72 amino acids  =  7.1 kDa
3 segments
   Segment 3:  
   414 .. 515  =  102 bp
   34 amino acids  =  3.4 kDa
Product: biotinylated sequence derived from the C-terminus of the α-subunit of Klebsiella pneumoniae oxaloacetate decarboxylase
contains a unique biotinylation site
BioEase™ tag
300 .. 515  =  216 bp
72 amino acids  =  7.1 kDa
3 segments
Product: biotinylated sequence derived from the C-terminus of the α-subunit of Klebsiella pneumoniae oxaloacetate decarboxylase
contains a unique biotinylation site
Xpress™ tag
528 .. 551  =  24 bp
8 amino acids  =  998.0 Da
2 segments
   Segment 1:  
   528 .. 536  =  9 bp
   3 amino acids  =  409.4 Da
Product: Xpress™ epitope tag, including an enterokinase recognition and cleavage site
Xpress™ tag
528 .. 551  =  24 bp
8 amino acids  =  998.0 Da
2 segments
   Segment 2:  enterokinase site  
   537 .. 551  =  15 bp
   5 amino acids  =  606.5 Da
Product: Xpress™ epitope tag, including an enterokinase recognition and cleavage site
Xpress™ tag
528 .. 551  =  24 bp
8 amino acids  =  998.0 Da
2 segments
Product: Xpress™ epitope tag, including an enterokinase recognition and cleavage site
rop
4855 .. 5046  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein, which maintains plasmids at low copy number
rop
4855 .. 5046  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein, which maintains plasmids at low copy number
attR1
561 .. 685  =  125 bp
recombination site for the Gateway® LR reaction
attR1
561 .. 685  =  125 bp
recombination site for the Gateway® LR reaction
attR2
2120 .. 2244  =  125 bp
recombination site for the Gateway® LR reaction
attR2
2120 .. 2244  =  125 bp
recombination site for the Gateway® LR reaction
AmpR promoter
2701 .. 2805  =  105 bp
AmpR promoter
2701 .. 2805  =  105 bp
lacI promoter
7441 .. 7518  =  78 bp
lacI promoter
7441 .. 7518  =  78 bp
T7 terminator
2330 .. 2377  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
T7 terminator
2330 .. 2377  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
lac UV5 promoter
710 .. 740  =  31 bp
3 segments
   Segment 1:  -35  
   710 .. 715  =  6 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
710 .. 740  =  31 bp
3 segments
   Segment 2:  
   716 .. 733  =  18 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
710 .. 740  =  31 bp
3 segments
   Segment 3:  -10  
   734 .. 740  =  7 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
710 .. 740  =  31 bp
3 segments
E. coli lac promoter with an "up" mutation
lac operator
228 .. 252  =  25 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
lac operator
228 .. 252  =  25 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
T7 promoter
209 .. 227  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
209 .. 227  =  19 bp
promoter for bacteriophage T7 RNA polymerase
RBS
282 .. 288  =  7 bp
ribosome binding site
RBS
282 .. 288  =  7 bp
ribosome binding site
ORF:  1774 .. 2079  =  306 bp
ORF:  101 amino acids  =  11.7 kDa
ORF:  2806 .. 3666  =  861 bp
ORF:  286 amino acids  =  31.5 kDa
ORF:  794 .. 1453  =  660 bp
ORF:  219 amino acids  =  25.7 kDa
ORF:  5078 .. 5446  =  369 bp
ORF:  122 amino acids  =  14.2 kDa
ORF:  7616 .. 237  =  240 bp
ORF:  79 amino acids  =  8.0 kDa
ORF:  297 .. 596  =  300 bp
ORF:  99 amino acids  =  10.1 kDa
ORF:  5880 .. 6575  =  696 bp
ORF:  231 amino acids  =  25.4 kDa
ORF:  7479 .. 124  =  264 bp
ORF:  87 amino acids  =  9.5 kDa
ORF:  6338 .. 6601  =  264 bp
ORF:  87 amino acids  =  8.9 kDa
ORF:  7190 .. 72  =  501 bp
ORF:  166 amino acids  =  17.5 kDa
ORF:  4855 .. 5079  =  225 bp
ORF:  74 amino acids  =  8.5 kDa
ORF:  5443 .. 5781  =  339 bp
ORF:  112 amino acids  =  12.6 kDa
ORF:  6358 .. 7317  =  960 bp
ORF:  319 amino acids  =  34.1 kDa
ORF:  69 .. 296  =  228 bp
ORF:  75 amino acids  =  7.9 kDa
ORF:  3270 .. 3536  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  5685 .. 6302  =  618 bp
ORF:  205 amino acids  =  21.8 kDa
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Download pET104-DEST.dna file

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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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