pET-60-DEST

Gateway® Nova bacterial destination vector for expressing proteins tagged at the N-terminus with GST.

Sequence Author: MilliporeSigma (Novagen)

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BlpI (7397) AvrII (7379) 6xHis SalI (7218) PstI (7216) BfuAI - BspMI (7205) BmgBI (6995) SmaI - SrfI (6961) AvaI - BsoBI - TspMI - XmaI (6959) BbvCI (6815) PasI (6322) EcoRI (6090) lac UV5 promoter EagI - NotI (5770) PmlI (5632) thrombin site SwaI (5384) BstBI (5354) RBS XbaI (4916) T7 promoter BspDI * - ClaI * (4849) SgrAI (4805) BtgI (4687) SphI (4657) BstAPI (4448) BstEII (3942) ApaI (3921) PspOMI (3917) T7 terminator PsiI (370) AhdI (655) BsaI (716) BglI (775) FspI (877) PvuI (1025) PflFI - Tth111I (2518) PpuMI (3255) HpaI (3622) pET-60-DEST™ 7480 bp
BlpI  (7397)
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.
AvrII  (7379)
1 site
C C T A G G G G A T C C
SalI  (7218)
1 site
G T C G A C C A G C T G
PstI  (7216)
1 site
C T G C A G G A C G T C
BfuAI  (7205)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BfuAI recognition sequence.
Sticky ends from different BfuAI sites may not be compatible.
BfuAI is typically used at 50°C, but is 50% active at 37°C.
BspMI  (7205)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BspMI recognition sequence.
Sticky ends from different BspMI sites may not be compatible.
BmgBI  (6995)
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.
SmaI  (6961)
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  (6961)
1 site
G C C C G G G C C G G G C C C G
AvaI  (6959)
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  (6959)
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  (6959)
1 site
C C C G G G G G G C C C
XmaI  (6959)
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.
BbvCI  (6815)
1 site
C C T C A G C G G A G T C G
PasI  (6322)
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.
EcoRI  (6090)
1 site
G A A T T C C T T A A G
EagI  (5770)
1 site
C G G C C G G C C G G C
NotI  (5770)
1 site
G C G G C C G C C G C C G G C G
PmlI  (5632)
1 site
C A C G T G G T G C A C
SwaI  (5384)
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.
BstBI  (5354)
1 site
T T C G A A A A G C T T
XbaI  (4916)
1 site
T C T A G A A G A T C T
BspDI  (4849)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (4849)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
SgrAI  (4805)
1 site
C R C C G G Y G G Y G G C C R C

Efficient cleavage requires at least two copies of the SgrAI recognition sequence.
BtgI  (4687)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
SphI  (4657)
1 site
G C A T G C C G T A C G
BstAPI  (4448)
1 site
G C A N N N N N T G C C G T N N N N N A C G

Sticky ends from different BstAPI sites may not be compatible.
BstEII  (3942)
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.
ApaI  (3921)
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  (3917)
1 site
G G G C C C C C C G G G
PsiI  (370)
1 site
T T A T A A A A T A T T
AhdI  (655)
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.
BsaI  (716)
1 site
G G T C T C N C C A G A G N ( N ) 4

Sticky ends from different BsaI sites may not be compatible.
BsaI can be used between 37°C and 50°C.
BglI  (775)
1 site
G C C N N N N N G G C C G G N N N N N C C G

Sticky ends from different BglI sites may not be compatible.
FspI  (877)
1 site
T G C G C A A C G C G T
PvuI  (1025)
1 site
C G A T C G G C T A G C
PflFI  (2518)
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  (2518)
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.
PpuMI  (3255)
1 site
R G G W C C Y Y C C W G G R

Sticky ends from different PpuMI sites may not be compatible.
HpaI  (3622)
1 site
G T T A A C C A A T T G
lacI
3397 .. 4479  =  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
3397 .. 4479  =  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
582 .. 1442  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   582 .. 1373  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
582 .. 1442  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   1374 .. 1442  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
582 .. 1442  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
GST
4957 .. 5610  =  654 bp
218 amino acids  =  25.5 kDa
Product: glutathione S-transferase from Schistosoma japonicum
GST
4957 .. 5610  =  654 bp
218 amino acids  =  25.5 kDa
Product: glutathione S-transferase from Schistosoma japonicum
thrombin site
5623 .. 5640  =  18 bp
6 amino acids  =  627.7 Da
Product: thrombin recognition and cleavage site
thrombin site
5623 .. 5640  =  18 bp
6 amino acids  =  627.7 Da
Product: thrombin recognition and cleavage site
CmR
5877 .. 6536  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
CmR
5877 .. 6536  =  660 bp
219 amino acids  =  25.7 kDa
Product: chloramphenicol acetyltransferase
confers resistance to chloramphenicol
ori
1616 .. 2204  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
1616 .. 2204  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
12 .. 467  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
12 .. 467  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
ccdB
6878 .. 7183  =  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
6878 .. 7183  =  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.
rop
2634 .. 2825  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein, which maintains plasmids at low copy number
rop
2634 .. 2825  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein, which maintains plasmids at low copy number
attR1
5644 .. 5768  =  125 bp
recombination site for the Gateway® LR reaction
attR1
5644 .. 5768  =  125 bp
recombination site for the Gateway® LR reaction
attR2
7224 .. 7348  =  125 bp
recombination site for the Gateway® LR reaction
attR2
7224 .. 7348  =  125 bp
recombination site for the Gateway® LR reaction
AmpR promoter
1443 .. 1535  =  93 bp
AmpR promoter
1443 .. 1535  =  93 bp
lacI promoter
4480 .. 4557  =  78 bp
lacI promoter
4480 .. 4557  =  78 bp
T7 terminator
7408 .. 7455  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
T7 terminator
7408 .. 7455  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
lac UV5 promoter
5793 .. 5823  =  31 bp
3 segments
   Segment 1:  -35  
   5793 .. 5798  =  6 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
5793 .. 5823  =  31 bp
3 segments
   Segment 2:  
   5799 .. 5816  =  18 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
5793 .. 5823  =  31 bp
3 segments
   Segment 3:  -10  
   5817 .. 5823  =  7 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
5793 .. 5823  =  31 bp
3 segments
E. coli lac promoter with an "up" mutation
lac operator
4889 .. 4913  =  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
4889 .. 4913  =  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
4870 .. 4888  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
4870 .. 4888  =  19 bp
promoter for bacteriophage T7 RNA polymerase
6xHis
7350 .. 7367  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
7350 .. 7367  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
RBS
4944 .. 4950  =  7 bp
ribosome binding site
RBS
4944 .. 4950  =  7 bp
ribosome binding site
ORF:  712 .. 978  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  2857 .. 3225  =  369 bp
ORF:  122 amino acids  =  14.2 kDa
ORF:  4957 .. 5679  =  723 bp
ORF:  240 amino acids  =  27.9 kDa
ORF:  4655 .. 4906  =  252 bp
ORF:  83 amino acids  =  8.6 kDa
ORF:  6878 .. 7183  =  306 bp
ORF:  101 amino acids  =  11.7 kDa
ORF:  3363 .. 3614  =  252 bp
ORF:  83 amino acids  =  9.1 kDa
ORF:  4518 .. 4781  =  264 bp
ORF:  87 amino acids  =  9.5 kDa
ORF:  5877 .. 6536  =  660 bp
ORF:  219 amino acids  =  25.7 kDa
ORF:  3377 .. 3640  =  264 bp
ORF:  87 amino acids  =  8.9 kDa
ORF:  4229 .. 4729  =  501 bp
ORF:  166 amino acids  =  17.5 kDa
ORF:  3397 .. 4356  =  960 bp
ORF:  319 amino acids  =  34.1 kDa
ORF:  582 .. 1442  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
ORF:  2634 .. 2858  =  225 bp
ORF:  74 amino acids  =  8.5 kDa