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Plasmid Files

pJET1.2

Positive selection cloning vector with a lethal insert that allows for efficient recovery of blunt-ended PCR products.

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pJET1.2 Sequence and MappJET1.2.dna
Map and Sequence File   
Sequence Author:  Thermo Scientific (Fermentas)
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 PstI (9) XmnI (2594) ScaI (2475) TatI (2473) PvuI (2365) FspI (2217) NmeAIII (2143) BglI (2115) BsrFI (2075) BpmI (2065) BsaI (2056) BmrI (2035) AhdI (1995) AlwNI (1518) BbsI (51) DraIII (125) BfuAI - BspMI (210) pJET1.2 forward sequencing primer (310 .. 332) EagI - NotI (329) BglII (337) BspEI (343) AvaI - BsoBI - PaeR7I - PspXI - XhoI (352) BmeT110I (353) EcoRV (371) XbaI (377) BglII (383) BtgI - NcoI - StyI (408) BspDI - ClaI (418) pJET1.2 reverse sequencing primer (405 .. 428) HindIII (624) BbvCI - Bpu10I (718) BsmI (722) BsmBI (745) PmeI (764) MfeI (892) BspQI - SapI (986) DrdI (1210) BseYI (1406) PspFI (1410) pJET1.2 2974 bp
PstI  (9)
1 site
C T G C A G G A C G T C
XmnI  (2594)
1 site
G A A N N N N T T C C T T N N N N A A G
ScaI  (2475)
1 site
A G T A C T T C A T G A
TatI  (2473)
1 site
W G T A C W W C A T G W
PvuI  (2365)
1 site
C G A T C G G C T A G C
FspI  (2217)
1 site
T G C G C A A C G C G T
NmeAIII  (2143)
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  (2115)
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.
BsrFI  (2075)
1 site
R C C G G Y Y G G C C R

Efficient cleavage requires at least two copies of the BsrFI
recognition sequence.
After cleavage, BsrFI can remain bound to DNA and alter its
electrophoretic mobility.
BpmI  (2065)
1 site
C T G G A G ( N ) 14 N N G A C C T C ( N ) 14

Efficient cleavage requires at least two copies of the BpmI
recognition sequence.
Sticky ends from different BpmI sites may not be compatible.
After cleavage, BpmI can remain bound to DNA and alter its
electrophoretic mobility.
BpmI quickly loses activity at 37°C.
BsaI  (2056)
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.
BmrI  (2035)
1 site
A C T G G G ( N ) 4 N T G A C C C ( N ) 4

The 1-base overhangs produced by BmrI may be hard to ligate.
Sticky ends from different BmrI sites may not be compatible.
Unlike most restriction enzymes, BmrI can cleave DNA in the
absence of magnesium.
AhdI  (1995)
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.
AlwNI  (1518)
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.
BbsI  (51)
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.
DraIII  (125)
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.
BfuAI  (210)
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  (210)
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.
EagI  (329)
1 site
C G G C C G G C C G G C
NotI  (329)
1 site
G C G G C C G C C G C C G G C G
BglII  (337)
2 sites
A G A T C T T C T A G A
BspEI  (343)
1 site
T C C G G A A G G C C T
AvaI  (352)
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  (352)
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.
PaeR7I  (352)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (352)
1 site
V C T C G A G B B G A G C T C V
XhoI  (352)
1 site
C T C G A G G A G C T C
BmeT110I  (353)
1 site
C Y C G R G G R G C Y C
EcoRV  (371)
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.
XbaI  (377)
1 site
T C T A G A A G A T C T
BglII  (383)
2 sites
A G A T C T T C T A G A
BtgI  (408)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
NcoI  (408)
1 site
C C A T G G G G T A C C
StyI  (408)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
BspDI  (418)
1 site
A T C G A T T A G C T A
ClaI  (418)
1 site
A T C G A T T A G C T A
HindIII  (624)
1 site
A A G C T T T T C G A A
BbvCI  (718)
1 site
C C T C A G C G G A G T C G
Bpu10I  (718)
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.
BsmI  (722)
1 site
G A A T G C N C T T A C G N

Sticky ends from different BsmI sites may not be compatible.
BsmBI  (745)
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.
PmeI  (764)
1 site
G T T T A A A C C A A A T T T G
MfeI  (892)
1 site
C A A T T G G T T A A C
BspQI  (986)
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  (986)
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.
DrdI  (1210)
1 site
G A C N N N N N N G T C C T G N N N N N N C A G

Sticky ends from different DrdI sites may not be compatible.
BseYI  (1406)
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  (1410)
1 site
C C C A G C G G G T C G
pJET1.2 forward sequencing primer
23-mer  /  61% GC
1 binding site
310 .. 332  =  23 annealed bases
Tm  =  62°C
pJET1.2 reverse sequencing primer
24-mer  /  42% GC
1 binding site
405 .. 428  =  24 annealed bases
Tm  =  58°C
AmpR
1922 .. 2782  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   1922 .. 2713  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1922 .. 2782  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   2714 .. 2782  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1922 .. 2782  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
Eco47I/T7
16 .. 753  =  738 bp
245 amino acids  =  28.5 kDa
Product: mutant version of the Eco47I restriction
endonuclease
cleaves double-stranded DNA at GGWCC sites in the
absence of protective methylation
Eco47I/T7
16 .. 753  =  738 bp
245 amino acids  =  28.5 kDa
Product: mutant version of the Eco47I restriction
endonuclease
cleaves double-stranded DNA at GGWCC sites in the
absence of protective methylation
ori
1163 .. 1751  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
1163 .. 1751  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
AmpR promoter
2783 .. 2887  =  105 bp
AmpR promoter
2783 .. 2887  =  105 bp
lac UV5 promoter
806 .. 836  =  31 bp
   Segment 3:  -10  
   806 .. 812  =  7 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
806 .. 836  =  31 bp
   Segment 2:  
   813 .. 830  =  18 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
806 .. 836  =  31 bp
   Segment 1:  -35  
   831 .. 836  =  6 bp
E. coli lac promoter with an "up" mutation
lac UV5 promoter
806 .. 836  =  31 bp
3 segments
E. coli lac promoter with an "up" mutation
lac operator
782 .. 798  =  17 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
782 .. 798  =  17 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).
MCS
328 .. 422  =  95 bp
multiple cloning site with a central Eco32I (EcoRV)
site
MCS
328 .. 422  =  95 bp
multiple cloning site with a central Eco32I (EcoRV)
site
T7 promoter
305 .. 323  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
305 .. 323  =  19 bp
promoter for bacteriophage T7 RNA polymerase
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