pQE-T7-2

Vector for expressing C-terminally 10xHis-tagged proteins in bacteria using the QIAgenes system.

Sequence Author: Qiagen

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StyI (57) DraIII (5049) PsiI (4921) AsiSI - PvuI (4349) SmaI (4223) TspMI - XmaI (4221) BspDI - ClaI (4040) NruI (4006) AcuI (3695) AlwNI (3563) BssSI - BssSαI (3320) PciI (3147) BspQI - SapI (3031) TatI (2954) BstZ17I (2921) AccI (2920) PflFI - Tth111I (2895) BlpI (80) stop codons 10xHis PaeR7I - PspXI - XhoI (173) PstI (185) BmgBI (188) Acc65I (192) KpnI (196) EagI - NotI - SacII (200) Eco53kI (211) SacI (213) NdeI (220) RBS XbaI (261) lac operator T7 promoter BglII (327) SgrAI (368) SphI (524) BstAPI (732) MluI (1049) BclI * (1063) BstEII (1230) NmeAIII (1255) PspOMI (1256) ApaI (1260) BssHII (1460) EcoRV (1499) HincII - HpaI (1555) PshAI (1894) BglI (2113) FspI - FspAI (2131) PpuMI (2156) pQE-T7-2 5291 bp
StyI  (57)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
DraIII  (5049)
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.
PsiI  (4921)
1 site
T T A T A A A A T A T T
AsiSI  (4349)
1 site
G C G A T C G C C G C T A G C G
PvuI  (4349)
1 site
C G A T C G G C T A G C
SmaI  (4223)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
TspMI  (4221)
1 site
C C C G G G G G G C C C
XmaI  (4221)
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.
BspDI  (4040)
1 site
A T C G A T T A G C T A
ClaI  (4040)
1 site
A T C G A T T A G C T A
NruI  (4006)
1 site
T C G C G A A G C G C T
AcuI  (3695)
1 site
C T G A A G ( N ) 14 N N G A C T T C ( N ) 14

Cleavage may be enhanced when more than one copy of the AcuI recognition sequence is present.
Sticky ends from different AcuI sites may not be compatible.
After cleavage, AcuI can remain bound to DNA and alter its electrophoretic mobility.
For full activity, add fresh S-adenosylmethionine (SAM).
AlwNI  (3563)
1 site
C A G N N N C T G G T C N N N G A C

Sticky ends from different AlwNI sites may not be compatible.
BssSI  (3320)
1 site
C A C G A G G T G C T C
BssSαI  (3320)
1 site
C A C G A G G T G C T C
PciI  (3147)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspQI  (3031)
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  (3031)
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.
TatI  (2954)
1 site
W G T A C W W C A T G W
BstZ17I  (2921)
1 site
G T A T A C C A T A T G
AccI  (2920)
1 site
G T M K A C C A K M T G

Efficient cleavage with AccI requires ≥13 bp on each side of the recognition sequence.
Sticky ends from different AccI sites may not be compatible.
PflFI  (2895)
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  (2895)
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.
BlpI  (80)
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.
PaeR7I  (173)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (173)
1 site
V C T C G A G B B G A G C T C V
XhoI  (173)
1 site
C T C G A G G A G C T C
PstI  (185)
1 site
C T G C A G G A C G T C
BmgBI  (188)
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.
Acc65I  (192)
1 site
G G T A C C C C A T G G
KpnI  (196)
1 site
G G T A C C C C A T G G
EagI  (200)
1 site
C G G C C G G C C G G C
NotI  (200)
1 site
G C G G C C G C C G C C G G C G
SacII  (200)
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.
Eco53kI  (211)
1 site
G A G C T C C T C G A G
SacI  (213)
1 site
G A G C T C C T C G A G
NdeI  (220)
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.
XbaI  (261)
1 site
T C T A G A A G A T C T
BglII  (327)
1 site
A G A T C T T C T A G A
SgrAI  (368)
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.
SphI  (524)
1 site
G C A T G C C G T A C G
BstAPI  (732)
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.
MluI  (1049)
1 site
A C G C G T T G C G C A
BclI  (1063)
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.
BstEII  (1230)
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.
NmeAIII  (1255)
1 site
G C C G A G ( N ) 18-19 N N C G G C T C ( N ) 18-19

Efficient cleavage requires at least two copies of the NmeAIII recognition sequence.
Sticky ends from different NmeAIII sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
PspOMI  (1256)
1 site
G G G C C C C C C G G G
ApaI  (1260)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
BssHII  (1460)
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.
EcoRV  (1499)
1 site
G A T A T C C T A T A G

EcoRV is reportedly more prone than its isoschizomer Eco32I to delete a base after cleavage.
HincII  (1555)
1 site
G T Y R A C C A R Y T G
HpaI  (1555)
1 site
G T T A A C C A A T T G
PshAI  (1894)
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.
BglI  (2113)
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  (2131)
1 site
T G C G C A A C G C G T
FspAI  (2131)
1 site
R T G C G C A Y Y A C G C G T R
PpuMI  (2156)
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.
lacI
699 .. 1781  =  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
699 .. 1781  =  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).
KanR
3918 .. 4733  =  816 bp
271 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin
KanR
3918 .. 4733  =  816 bp
271 amino acids  =  31.0 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin
ori
3208 .. 3796  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
3208 .. 3796  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
f1 ori
4825 .. 5280  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
4825 .. 5280  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
rop
2590 .. 2781  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein
rop
2590 .. 2781  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein
lacI promoter
621 .. 698  =  78 bp
lacI promoter
621 .. 698  =  78 bp
MCS
173 .. 224  =  52 bp
multiple cloning site
MCS
173 .. 224  =  52 bp
multiple cloning site
T7 terminator
26 .. 73  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
T7 terminator
26 .. 73  =  48 bp
transcription terminator for bacteriophage T7 RNA polymerase
10xHis
143 .. 172  =  30 bp
10 amino acids  =  1.4 kDa
Product: 10xHis affinity tag
10xHis
143 .. 172  =  30 bp
10 amino acids  =  1.4 kDa
Product: 10xHis affinity tag
lac operator
269 .. 293  =  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
269 .. 293  =  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
294 .. 312  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
294 .. 312  =  19 bp
promoter for bacteriophage T7 RNA polymerase
stop codons
137 .. 142  =  6 bp
two tandem stop codons
stop codons
137 .. 142  =  6 bp
two tandem stop codons
RBS
232 .. 237  =  6 bp
ribosome binding site
RBS
232 .. 237  =  6 bp
ribosome binding site
ATG
219 .. 221  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
219 .. 221  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ORF:  1837 .. 2193  =  357 bp
ORF:  118 amino acids  =  13.0 kDa
ORF:  2557 .. 2781  =  225 bp
ORF:  74 amino acids  =  8.5 kDa
ORF:  449 .. 949  =  501 bp
ORF:  166 amino acids  =  17.5 kDa
ORF:  1538 .. 1801  =  264 bp
ORF:  87 amino acids  =  8.9 kDa
ORF:  207 .. 452  =  246 bp
ORF:  81 amino acids  =  8.6 kDa
ORF:  822 .. 1781  =  960 bp
ORF:  319 amino acids  =  34.1 kDa
ORF:  3918 .. 4733  =  816 bp
ORF:  271 amino acids  =  31.0 kDa
ORF:  2190 .. 2558  =  369 bp
ORF:  122 amino acids  =  14.2 kDa
ORF:  284 .. 523  =  240 bp
ORF:  79 amino acids  =  8.0 kDa
ORF:  397 .. 660  =  264 bp
ORF:  87 amino acids  =  9.5 kDa
ORF:  1564 .. 1815  =  252 bp
ORF:  83 amino acids  =  9.1 kDa
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