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Plasmid Files

pENTR™/GeneBLAzer®

Gateway® entry clone to be used as a control for fluorescence detection of β-lactamase with the GeneBLAzer® system.

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pENTR GeneBLAzer Sequence and MappENTR GeneBLAzer.dna
Map and Sequence File   
Sequence Author:  Invitrogen (Life Technologies)
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 PciI (3333) DrdI (3231) BciVI (3135) PspFI (3033) BseYI (3029) AlwNI (2924) PflMI (2421) Bpu10I (2175) BsmBI (2174) AsiSI (2158) SspI (2083) EcoNI (2070) NruI (1815) NspI (3337) BspQI - SapI (111) BbsI (437) HincII - HpaI (501) AflII (554) AvaI - BsoBI (560) BmeT110I (561) PspOMI (563) EcoO109I (564) ApaI (567) BtgI - NcoI - StyI (669) XmnI (792) DraI (814) ScaI (911) FspI (1169) NmeAIII (1245) BglI (1274) BpmI (1323) BsaI (1326) AccI (1463) EcoRV (1576) pENTR™/GeneBLAzer® 3339 bp
PciI  (3333)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
DrdI  (3231)
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.
BciVI  (3135)
1 site
G T A T C C ( N ) 5 N C A T A G G ( N ) 5

The 1-base overhangs produced by BciVI may be hard to ligate.
Sticky ends from different BciVI sites may not be compatible.
PspFI  (3033)
1 site
C C C A G C G G G T C G
BseYI  (3029)
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.
AlwNI  (2924)
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.
PflMI  (2421)
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.
Bpu10I  (2175)
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.
BsmBI  (2174)
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.
AsiSI  (2158)
1 site
G C G A T C G C C G C T A G C G
SspI  (2083)
1 site
A A T A T T T T A T A A
EcoNI  (2070)
1 site
C C T N N N N N A G G G G A N N N N N T C C

The 1-base overhangs produced by EcoNI may be hard to ligate.
Sticky ends from different EcoNI sites may not be compatible.
NruI  (1815)
1 site
T C G C G A A G C G C T
NspI  (3337)
1 site
R C A T G Y Y G T A C R
BspQI  (111)
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  (111)
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.
BbsI  (437)
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.
HincII  (501)
1 site
G T Y R A C C A R Y T G
HpaI  (501)
1 site
G T T A A C C A A T T G
AflII  (554)
1 site
C T T A A G G A A T T C

The sticky ends produced by AflII are hard to ligate.
AvaI  (560)
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  (560)
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.
BmeT110I  (561)
1 site
C Y C G R G G R G C Y C
PspOMI  (563)
1 site
G G G C C C C C C G G G
EcoO109I  (564)
1 site
R G G N C C Y Y C C N G G R

Sticky ends from different EcoO109I sites may not be compatible.
ApaI  (567)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
BtgI  (669)
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  (669)
1 site
C C A T G G G G T A C C
StyI  (669)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
XmnI  (792)
1 site
G A A N N N N T T C C T T N N N N A A G
DraI  (814)
1 site
T T T A A A A A A T T T
ScaI  (911)
1 site
A G T A C T T C A T G A
FspI  (1169)
1 site
T G C G C A A C G C G T
NmeAIII  (1245)
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  (1274)
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.
BpmI  (1323)
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  (1326)
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.
AccI  (1463)
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.
EcoRV  (1576)
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.
KanR
1733 .. 2542  =  810 bp
269 amino acids  =  30.8 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418
(Geneticin®) in eukaryotes
KanR
1733 .. 2542  =  810 bp
269 amino acids  =  30.8 kDa
Product: aminoglycoside phosphotransferase
confers resistance to kanamycin in bacteria or G418
(Geneticin®) in eukaryotes
bla(M)
671 .. 1465  =  795 bp
264 amino acids  =  29.1 kDa
Product: β-lactamase lacking the signal sequence
allows cytosolic expression of β-lactamase
bla(M)
671 .. 1465  =  795 bp
264 amino acids  =  29.1 kDa
Product: β-lactamase lacking the signal sequence
allows cytosolic expression of β-lactamase
ori
2689 .. 3277  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
2689 .. 3277  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
attL1
569 .. 667  =  99 bp
recombination site for the Gateway® LR reaction
attL1
569 .. 667  =  99 bp
recombination site for the Gateway® LR reaction
rrnB T1 terminator
387 .. 473  =  87 bp
transcription terminator T1 from the E. coli rrnB
gene
rrnB T1 terminator
387 .. 473  =  87 bp
transcription terminator T1 from the E. coli rrnB
gene
rrnB T2 terminator
268 .. 295  =  28 bp
transcription terminator T2 from the E. coli rrnB
gene
rrnB T2 terminator
268 .. 295  =  28 bp
transcription terminator T2 from the E. coli rrnB
gene
attL2
1464 .. 1563  =  100 bp
recombination site for the Gateway® LR reaction
attL2
1464 .. 1563  =  100 bp
recombination site for the Gateway® LR reaction
Kozak sequence
668 .. 674  =  7 bp
translation initiation signal for mammalian cells
Kozak sequence
668 .. 674  =  7 bp
translation initiation signal for mammalian cells
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