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Plasmid Files

pET-52b(+)

Bacterial vector for expressing proteins with a cleavable N-terminal Strep-Tag II tag and a cleavable C-terminal 10xHis tag.

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pET-52b(+).dna
Map and Sequence File:    Download    Open   
Sequence Author:  Novagen (EMD Millipore)
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 thrombin site 10xHis AvrII (97) BlpI (80) DraIII (4985) PsiI (4857) SspI (4777) AhdI (4573) BsaI (4507) BglI (4455) FspI (4350) PvuI (4204) ScaI (4092) AlwNI (3379) PciI (2963) BspQI - SapI (2847) BstZ17I (2734) PflFI - Tth111I (2708) Eco53kI (169) SacI (171) EagI - NotI (175) SalI (182) BsrGI (189) BamHI (195) Acc65I (202) AvaI - BsoBI - KpnI - TspMI - XmaI (206) BmeT110I (207) SmaI (208) KflI (210) HRV 3C site BstBI (243) Strep-Tag II NcoI (268) ATG XbaI (307) lac operator T7 promoter BspDI * - ClaI * (376) SgrAI (418) SphI (574) PflMI (681) BstAPI (782) MluI (1099) BclI * (1113) BstEII (1280) PspOMI (1306) ApaI (1310) BssHII (1510) HpaI (1605) Bpu10I (2069) pET-52b(+) 5227 bp
AvrII  (97)
1 site
C C T A G G G G A T C C
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.
DraIII  (4985)
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  (4857)
1 site
T T A T A A A A T A T T
SspI  (4777)
1 site
A A T A T T T T A T A A
AhdI  (4573)
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  (4507)
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  (4455)
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  (4350)
1 site
T G C G C A A C G C G T
PvuI  (4204)
1 site
C G A T C G G C T A G C
ScaI  (4092)
1 site
A G T A C T T C A T G A
AlwNI  (3379)
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.
PciI  (2963)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspQI  (2847)
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  (2847)
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.
BstZ17I  (2734)
1 site
G T A T A C C A T A T G
PflFI  (2708)
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  (2708)
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.
Eco53kI  (169)
1 site
G A G C T C C T C G A G
SacI  (171)
1 site
G A G C T C C T C G A G
EagI  (175)
1 site
C G G C C G G C C G G C
NotI  (175)
1 site
G C G G C C G C C G C C G G C G
SalI  (182)
1 site
G T C G A C C A G C T G
BsrGI  (189)
1 site
T G T A C A A C A T G T

BsrGI is typically used at 37°C, but is even more active at 60°C.
BamHI  (195)
1 site
G G A T C C C C T A G G

After cleavage, BamHI-HF™ (but not the original BamHI) can
remain bound to DNA and alter its electrophoretic mobility.
Acc65I  (202)
1 site
G G T A C C C C A T G G
AvaI  (206)
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  (206)
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.
KpnI  (206)
1 site
G G T A C C C C A T G G
TspMI  (206)
1 site
C C C G G G G G G C C C
XmaI  (206)
1 site
C C C G G G G G G C C C

Efficient cleavage requires at least two copies of the XmaI
recognition sequence.
Full cleavage with XmaI may require a long incubation.
BmeT110I  (207)
1 site
C Y C G R G G R G C Y C
SmaI  (208)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
KflI  (210)
1 site
G G G W C C C C C C W G G G

Sticky ends from different KflI sites may not be compatible.
BstBI  (243)
1 site
T T C G A A A A G C T T
NcoI  (268)
1 site
C C A T G G G G T A C C
XbaI  (307)
1 site
T C T A G A A G A T C T
BspDI  (376)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (376)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
SgrAI  (418)
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  (574)
1 site
G C A T G C C G T A C G
PflMI  (681)
1 site
C C A N N N N N T G G G G T N N N N N A C C

Sticky ends from different PflMI sites may not be compatible.
BstAPI  (782)
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  (1099)
1 site
A C G C G T T G C G C A
BclI  (1113)
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  (1280)
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.
PspOMI  (1306)
1 site
G G G C C C C C C G G G
ApaI  (1310)
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  (1510)
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.
HpaI  (1605)
1 site
G T T A A C C A A T T G
Bpu10I  (2069)
1 site
C C T N A G C G G A N T C G

Efficient cleavage requires at least two copies of the Bpu10I
recognition sequence.
This recognition sequence is asymmetric, so ligating sticky ends
generated by Bpu10I will not always regenerate a Bpu10I site.
Sticky ends from different Bpu10I sites may not be compatible.
lacI
749 .. 1831  =  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
749 .. 1831  =  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
3786 .. 4646  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   3786 .. 3854  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3786 .. 4646  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   3855 .. 4646  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3786 .. 4646  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
3024 .. 3612  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3024 .. 3612  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
4761 .. 5216  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
4761 .. 5216  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
rop
2403 .. 2594  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein
rop
2403 .. 2594  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein
lacI promoter
671 .. 748  =  78 bp
lacI promoter
671 .. 748  =  78 bp
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
MCS
167 .. 208  =  42 bp
multiple cloning site
MCS
167 .. 208  =  42 bp
multiple cloning site
10xHis
110 .. 139  =  30 bp
10 amino acids  =  1.4 kDa
Product: 10xHis affinity tag
10xHis
110 .. 139  =  30 bp
10 amino acids  =  1.4 kDa
Product: 10xHis affinity tag
lac operator
315 .. 339  =  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
315 .. 339  =  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).
HRV 3C site
209 .. 232  =  24 bp
8 amino acids  =  902.1 Da
Product: recognition and cleavage site for human
rhinovirus 3C protease
HRV 3C site
209 .. 232  =  24 bp
8 amino acids  =  902.1 Da
Product: recognition and cleavage site for human
rhinovirus 3C protease
Strep-Tag II
239 .. 262  =  24 bp
8 amino acids  =  1.1 kDa
Product: peptide that binds Strep-Tactin, an
engineered form of streptavidin
Strep-Tag II
239 .. 262  =  24 bp
8 amino acids  =  1.1 kDa
Product: peptide that binds Strep-Tactin, an
engineered form of streptavidin
T7 promoter
340 .. 358  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
340 .. 358  =  19 bp
promoter for bacteriophage T7 RNA polymerase
thrombin site
146 .. 163  =  18 bp
6 amino acids  =  627.8 Da
Product: thrombin recognition and cleavage site
thrombin site
146 .. 163  =  18 bp
6 amino acids  =  627.8 Da
Product: thrombin recognition and cleavage site
RBS
279 .. 284  =  6 bp
ribosome binding site
RBS
279 .. 284  =  6 bp
ribosome binding site
ATG
269 .. 271  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
269 .. 271  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
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