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Plasmid Files

pET-23b(+)

Bacterial vector for expression of N-terminally T7-tagged proteins. For other reading frames, use pET‑23a(+) or pET‑23c(+).

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pET-23b(+).dna
Map and Sequence File:    Download    Open   
Sequence Author:  Novagen (EMD Millipore)
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 NaeI (3526) NgoMIV (3524) DraIII (3423) PsiI (3295) BsaHI (2817) ScaI (2760) PvuI (2650) PstI (2525) NmeAIII (2428) BglI (2400) AhdI (2280) StyI (57) BlpI (80) 6xHis PaeR7I - PspXI - XhoI (158) EagI - NotI (166) HindIII (173) SalI (179) MCS HincII (181) Eco53kI (188) SacI (190) EcoRI (192) BamHI (198) T7 tag NheI (230) BmtI (234) NdeI (237) RBS XbaI (275) BglII (333) MscI * (358) BtgI (359) FspAI (368) Bpu10I (493) BbsI (505) BsgI * (547) AfeI (641) PvuII (978) BsmBI (1028) PfoI (1029) PflFI - Tth111I (1132) BstZ17I (1158) BspQI - SapI (1271) AflIII - PciI (1387) BseYI (1691) PspFI (1695) AlwNI (1803) pET-23b(+) 3665 bp
NaeI  (3526)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NaeI
recognition sequence.
NgoMIV  (3524)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NgoMIV
recognition sequence.
DraIII  (3423)
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  (3295)
1 site
T T A T A A A A T A T T
BsaHI  (2817)
1 site
G R C G Y C C Y G C R G

BsaHI is typically used at 37°C, but is even more active at 60°C.
ScaI  (2760)
1 site
A G T A C T T C A T G A
PvuI  (2650)
1 site
C G A T C G G C T A G C
PstI  (2525)
1 site
C T G C A G G A C G T C
NmeAIII  (2428)
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).
BglI  (2400)
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.
AhdI  (2280)
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.
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.
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  (158)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (158)
1 site
V C T C G A G B B G A G C T C V
XhoI  (158)
1 site
C T C G A G G A G C T C
EagI  (166)
1 site
C G G C C G G C C G G C
NotI  (166)
1 site
G C G G C C G C C G C C G G C G
HindIII  (173)
1 site
A A G C T T T T C G A A
SalI  (179)
1 site
G T C G A C C A G C T G
HincII  (181)
1 site
G T Y R A C C A R Y T G
Eco53kI  (188)
1 site
G A G C T C C T C G A G
SacI  (190)
1 site
G A G C T C C T C G A G
EcoRI  (192)
1 site
G A A T T C C T T A A G
BamHI  (198)
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.
NheI  (230)
1 site
G C T A G C C G A T C G
BmtI  (234)
1 site
G C T A G C C G A T C G
NdeI  (237)
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  (275)
1 site
T C T A G A A G A T C T
BglII  (333)
1 site
A G A T C T T C T A G A
MscI  (358)
1 site
T G G C C A A C C G G T
* Blocked by Dcm methylation.
BtgI  (359)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
FspAI  (368)
1 site
R T G C G C A Y Y A C G C G T R
Bpu10I  (493)
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.
BbsI  (505)
1 site
G A A G A C N N C T T C T G N N ( N ) 4

Sticky ends from different BbsI sites may not be compatible.
BbsI gradually loses activity when stored at -20°C.
BsgI  (547)
1 site
G T G C A G ( N ) 14 N N C A C G T C ( N ) 14
* Blocked by EcoKI methylation.
Efficient cleavage requires at least two copies of the BsgI
recognition sequence.
Sticky ends from different BsgI sites may not be compatible.
For full activity, add fresh S-adenosylmethionine (SAM).
AfeI  (641)
1 site
A G C G C T T C G C G A
PvuII  (978)
1 site
C A G C T G G T C G A C
BsmBI  (1028)
1 site
C G T C T C N G C A G A G N ( N ) 4

Sticky ends from different BsmBI sites may not be compatible.
PfoI  (1029)
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.
PflFI  (1132)
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  (1132)
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.
BstZ17I  (1158)
1 site
G T A T A C C A T A T G
BspQI  (1271)
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  (1271)
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.
AflIII  (1387)
1 site
A C R Y G T T G Y R C A

Sticky ends from different AflIII sites may not be compatible.
PciI  (1387)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BseYI  (1691)
1 site
C C C A G C G G G T C G

After cleavage, BseYI can remain bound to DNA and alter its
electrophoretic mobility.
PspFI  (1695)
1 site
C C C A G C G G G T C G
AlwNI  (1803)
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.
AmpR
2207 .. 3067  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 2:  
   2207 .. 2998  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
2207 .. 3067  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 1:  signal sequence  
   2999 .. 3067  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
2207 .. 3067  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
1448 .. 2036  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
1448 .. 2036  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
f1 ori
3199 .. 3654  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
f1 ori
3199 .. 3654  =  456 bp
f1 bacteriophage origin of replication; arrow
indicates direction of (+) strand synthesis
rop
827 .. 1018  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein
rop
827 .. 1018  =  192 bp
63 amino acids  =  7.2 kDa
Product: Rop protein
AmpR promoter
3068 .. 3172  =  105 bp
AmpR promoter
3068 .. 3172  =  105 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
158 .. 203  =  46 bp
multiple cloning site
MCS
158 .. 203  =  46 bp
multiple cloning site
T7 tag
206 .. 238  =  33 bp
11 amino acids  =  1.1 kDa
Product: epitope tag from a T7 major capsid protein
T7 tag
206 .. 238  =  33 bp
11 amino acids  =  1.1 kDa
Product: epitope tag from a T7 major capsid protein
T7 promoter
300 .. 318  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
300 .. 318  =  19 bp
promoter for bacteriophage T7 RNA polymerase
6xHis
140 .. 157  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
6xHis
140 .. 157  =  18 bp
6 amino acids  =  840.9 Da
Product: 6xHis affinity tag
RBS
247 .. 252  =  6 bp
ribosome binding site
RBS
247 .. 252  =  6 bp
ribosome binding site
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