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Plasmid Files

pTRE-Tight-BI-AcGFP1

Vector for co-expressing AcGFP1 and another gene with the Tet-On® Advanced or Tet-Off® Advanced system.

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pTRE-Tight-BI-AcGFP1 Sequence and MappTRE-Tight-BI-AcGFP1.dna
Map and Sequence File   
Sequence Author:  Clontech
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 EcoRI (3454) PluTI (3440) SfoI (3438) NarI (3437) KasI (3436) MscI (3379) AleI (3272) BstEII (3265) Bpu10I (3257) Bsu36I (3181) PfoI * (3167) BssHII (3122) DraIII (3006) EcoNI (2827) StyI (2823) EcoO109I - PpuMI (2820) PflMI (2757) BtgI (2756) XbaI * (2728) AatII (2536) ZraI (2534) SspI (2418) EarI (2409) XmnI (2213) ScaI (2094) PvuI (1984) FspI (1836) AseI (1786) PaeR7I - PspXI - XhoI (1) Acc65I (335) KpnI - TspMI - XmaI (339) SmaI (341) BamHI (344) MluI (362) NheI (368) BmtI (372) EagI - NotI (375) BspDI - ClaI (383) HindIII (388) SalI (394) AccI (395) EcoRV (402) PciI (721) NspI (725) DrdI (829) BseYI (1025) PspFI (1029) AlwNI (1137) AhdI (1614) BmrI (1654) BsaI (1675) BglI (1734) pTRE-Tight-BI-AcGFP1 3533 bp
EcoRI  (3454)
1 site
G A A T T C C T T A A G
PluTI  (3440)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the PluTI
recognition sequence.
SfoI  (3438)
1 site
G G C G C C C C G C G G
NarI  (3437)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the NarI
recognition sequence.
KasI  (3436)
1 site
G G C G C C C C G C G G
MscI  (3379)
1 site
T G G C C A A C C G G T
AleI  (3272)
1 site
C A C N N N N G T G G T G N N N N C A C
BstEII  (3265)
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.
Bpu10I  (3257)
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.
Bsu36I  (3181)
1 site
C C T N A G G G G A N T C C

Sticky ends from different Bsu36I sites may not be compatible.
PfoI  (3167)
1 site
T C C N G G A A G G N C C T
* Blocked by Dcm methylation.
Sticky ends from different PfoI sites may not be compatible.
BssHII  (3122)
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.
DraIII  (3006)
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.
EcoNI  (2827)
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.
StyI  (2823)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
EcoO109I  (2820)
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.
PpuMI  (2820)
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.
PflMI  (2757)
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.
BtgI  (2756)
1 site
C C R Y G G G G Y R C C

Sticky ends from different BtgI sites may not be compatible.
XbaI  (2728)
1 site
T C T A G A A G A T C T
* Blocked by Dam methylation.
AatII  (2536)
1 site
G A C G T C C T G C A G
ZraI  (2534)
1 site
G A C G T C C T G C A G
SspI  (2418)
1 site
A A T A T T T T A T A A
EarI  (2409)
1 site
C T C T T C N G A G A A G N N N N

Efficient cleavage requires at least two copies of the EarI
recognition sequence.
Sticky ends from different EarI sites may not be compatible.
XmnI  (2213)
1 site
G A A N N N N T T C C T T N N N N A A G
ScaI  (2094)
1 site
A G T A C T T C A T G A
PvuI  (1984)
1 site
C G A T C G G C T A G C
FspI  (1836)
1 site
T G C G C A A C G C G T
AseI  (1786)
1 site
A T T A A T T A A T T A
PaeR7I  (1)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (1)
1 site
V C T C G A G B B G A G C T C V
XhoI  (1)
1 site
C T C G A G G A G C T C
Acc65I  (335)
1 site
G G T A C C C C A T G G
KpnI  (339)
1 site
G G T A C C C C A T G G
TspMI  (339)
1 site
C C C G G G G G G C C C
XmaI  (339)
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.
SmaI  (341)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
BamHI  (344)
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.
MluI  (362)
1 site
A C G C G T T G C G C A
NheI  (368)
1 site
G C T A G C C G A T C G
BmtI  (372)
1 site
G C T A G C C G A T C G
EagI  (375)
1 site
C G G C C G G C C G G C
NotI  (375)
1 site
G C G G C C G C C G C C G G C G
BspDI  (383)
1 site
A T C G A T T A G C T A
ClaI  (383)
1 site
A T C G A T T A G C T A
HindIII  (388)
1 site
A A G C T T T T C G A A
SalI  (394)
1 site
G T C G A C C A G C T G
AccI  (395)
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  (402)
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.
PciI  (721)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
NspI  (725)
1 site
R C A T G Y Y G T A C R
DrdI  (829)
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  (1025)
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  (1029)
1 site
C C C A G C G G G T C G
AlwNI  (1137)
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.
AhdI  (1614)
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.
BmrI  (1654)
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.
BsaI  (1675)
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  (1734)
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.
AmpR
1541 .. 2401  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 2:  
   1541 .. 2332  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1541 .. 2401  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 1:  signal sequence  
   2333 .. 2401  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1541 .. 2401  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AcGFP1
2734 .. 3453  =  720 bp
239 amino acids  =  26.9 kDa
Product: Aequorea coerulescens GFP
mammalian codon-optimized
AcGFP1
2734 .. 3453  =  720 bp
239 amino acids  =  26.9 kDa
Product: Aequorea coerulescens GFP
mammalian codon-optimized
ori
782 .. 1370  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
782 .. 1370  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
bidirectional TRE promoter
3464 .. 318  =  388 bp
Tet-responsive bidirectional promoter PTight-BI,
consisting of seven tet operator sequences flanked
on each side by the minimal CMV promoter
bidirectional TRE promoter
3464 .. 318  =  388 bp
Tet-responsive bidirectional promoter PTight-BI,
consisting of seven tet operator sequences flanked
on each side by the minimal CMV promoter
AmpR promoter
2402 .. 2506  =  105 bp
AmpR promoter
2402 .. 2506  =  105 bp
SV40 poly(A) signal
519 .. 600  =  82 bp
SV40 polyadenylation signal
SV40 poly(A) signal
519 .. 600  =  82 bp
SV40 polyadenylation signal
SV40 poly(A) signal
2539 .. 2620  =  82 bp
SV40 polyadenylation signal
SV40 poly(A) signal
2539 .. 2620  =  82 bp
SV40 polyadenylation signal
MCS
335 .. 405  =  71 bp
multiple cloning site
MCS
335 .. 405  =  71 bp
multiple cloning site
tet operator
12 .. 30  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
12 .. 30  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
48 .. 66  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
48 .. 66  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
83 .. 101  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
83 .. 101  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
119 .. 137  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
119 .. 137  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
155 .. 173  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
155 .. 173  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
190 .. 208  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
190 .. 208  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
226 .. 244  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
226 .. 244  =  19 bp
bacterial operator O2 for the tetR and tetA genes
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