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pTRE3G-BI-Het1

Mammalian vector for doxycycline-inducible dual expression of FKBP- and FRB-tagged proteins, allowing drug-inducible heterodimerization.

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pTRE3G-BI-Het1 Sequence and MappTRE3G-BI-Het1.dna
Map and Sequence File   
Sequence Author:  Clontech
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 KpnI (3506) Acc65I (3502) ATG NcoI - StyI (3498) EcoNI (3454) SphI (3387) MluI (3205) 3' MCS-2 PstI (3203) PshAI (3184) NdeI (3183) AatII (2984) ZraI (2982) SspI (2866) XmnI (2661) ScaI (2542) PvuI (2432) FspI (2284) AseI (2234) NmeAIII (2210) BglI (2182) BsaI (2123) AhdI (2062) EcoRI (3514) PaeR7I - PspXI - XhoI (70) HindIII (322) KasI (343) NarI (344) SfoI (345) PluTI (347) SalI (452) AccI (453) BglII (468) BamHI (480) ATG AarI (491) BsmBI (517) BbsI (779) BspDI * - ClaI * (817) EagI - NotI (825) MreI - NgoMIV - SgrAI (830) NaeI (832) EcoRV (850) PciI (1169) DrdI (1277) pTRE3G-BI-Het1 3524 bp
KpnI  (3506)
1 site
G G T A C C C C A T G G
Acc65I  (3502)
1 site
G G T A C C C C A T G G
NcoI  (3498)
1 site
C C A T G G G G T A C C
StyI  (3498)
1 site
C C W W G G G G W W C C

Sticky ends from different StyI sites may not be compatible.
EcoNI  (3454)
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.
SphI  (3387)
1 site
G C A T G C C G T A C G
MluI  (3205)
1 site
A C G C G T T G C G C A
PstI  (3203)
1 site
C T G C A G G A C G T C
PshAI  (3184)
1 site
G A C N N N N G T C C T G N N N N C A G

PshAI quickly loses activity at 37°C, but can be used at 25°C for
long incubations.
NdeI  (3183)
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.
AatII  (2984)
1 site
G A C G T C C T G C A G
ZraI  (2982)
1 site
G A C G T C C T G C A G
SspI  (2866)
1 site
A A T A T T T T A T A A
XmnI  (2661)
1 site
G A A N N N N T T C C T T N N N N A A G
ScaI  (2542)
1 site
A G T A C T T C A T G A
PvuI  (2432)
1 site
C G A T C G G C T A G C
FspI  (2284)
1 site
T G C G C A A C G C G T
AseI  (2234)
1 site
A T T A A T T A A T T A
NmeAIII  (2210)
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  (2182)
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.
BsaI  (2123)
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.
AhdI  (2062)
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.
EcoRI  (3514)
1 site
G A A T T C C T T A A G
PaeR7I  (70)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (70)
1 site
V C T C G A G B B G A G C T C V
XhoI  (70)
1 site
C T C G A G G A G C T C
HindIII  (322)
1 site
A A G C T T T T C G A A
KasI  (343)
1 site
G G C G C C C C G C G G
NarI  (344)
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.
SfoI  (345)
1 site
G G C G C C C C G C G G
PluTI  (347)
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.
SalI  (452)
1 site
G T C G A C C A G C T G
AccI  (453)
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.
BglII  (468)
1 site
A G A T C T T C T A G A
BamHI  (480)
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.
AarI  (491)
1 site
C A C C T G C ( N ) 4 G T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the AarI
recognition sequence.
Sticky ends from different AarI sites may not be compatible.
After cleavage, AarI can remain bound to DNA and alter its
electrophoretic mobility.
BsmBI  (517)
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.
BbsI  (779)
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.
BspDI  (817)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
ClaI  (817)
1 site
A T C G A T T A G C T A
* Blocked by Dam methylation.
EagI  (825)
1 site
C G G C C G G C C G G C
NotI  (825)
1 site
G C G G C C G C C G C C G G C G
MreI  (830)
1 site
C G C C G G C G G C G G C C G C
NgoMIV  (830)
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.
SgrAI  (830)
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.
NaeI  (832)
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.
EcoRV  (850)
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  (1169)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
DrdI  (1277)
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.
AmpR
1989 .. 2849  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 2:  
   1989 .. 2780  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1989 .. 2849  =  861 bp
286 amino acids  =  31.6 kDa
   Segment 1:  signal sequence  
   2781 .. 2849  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
1989 .. 2849  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
ori
1230 .. 1818  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
1230 .. 1818  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
TRE3G BI promoter
1 .. 451  =  451 bp
3rd-generation Tet-responsive bidirectional
promoter that can be activated by binding of
Tet-On® 3G
TRE3G BI promoter
1 .. 451  =  451 bp
3rd-generation Tet-responsive bidirectional
promoter that can be activated by binding of
Tet-On® 3G
FKBP (DmrA)
495 .. 815  =  321 bp
107 amino acids  =  11.8 kDa
Product: human FK506-binding protein FKBP12
FKBP (DmrA)
495 .. 815  =  321 bp
107 amino acids  =  11.8 kDa
Product: human FK506-binding protein FKBP12
FRB* (DmrC)
3211 .. 3489  =  279 bp
93 amino acids  =  11.3 kDa
Product: T2098L mutant of FKBP-rapamycin binding
domain of human FRAP
binds synthetic ligands such as AP21967
FRB* (DmrC)
3211 .. 3489  =  279 bp
93 amino acids  =  11.3 kDa
Product: T2098L mutant of FKBP-rapamycin binding
domain of human FRAP
binds synthetic ligands such as AP21967
AmpR promoter
2850 .. 2954  =  105 bp
AmpR promoter
2850 .. 2954  =  105 bp
SV40 poly(A) signal
967 .. 1048  =  82 bp
SV40 polyadenylation signal
SV40 poly(A) signal
967 .. 1048  =  82 bp
SV40 polyadenylation signal
SV40 poly(A) signal
2987 .. 3068  =  82 bp
SV40 polyadenylation signal
SV40 poly(A) signal
2987 .. 3068  =  82 bp
SV40 polyadenylation signal
5' MCS-1
452 .. 485  =  34 bp
multiple cloning site upstream of DmrA
5' MCS-1
452 .. 485  =  34 bp
multiple cloning site upstream of DmrA
5' MCS-2
3502 .. 3519  =  18 bp
multiple cloning site upstream of DmrC
5' MCS-2
3502 .. 3519  =  18 bp
multiple cloning site upstream of DmrC
3' MCS-1
816 .. 831  =  16 bp
multiple cloning site downstream of DmrA
3' MCS-1
816 .. 831  =  16 bp
multiple cloning site downstream of DmrA
3' MCS-2
3199 .. 3210  =  12 bp
multiple cloning site downstream of DmrC
3' MCS-2
3199 .. 3210  =  12 bp
multiple cloning site downstream of DmrC
ATG
486 .. 488  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
486 .. 488  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
3499 .. 3501  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
3499 .. 3501  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
tet operator
81 .. 99  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
81 .. 99  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
117 .. 135  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
117 .. 135  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
153 .. 171  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
153 .. 171  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
189 .. 207  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
189 .. 207  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
225 .. 243  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
225 .. 243  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
261 .. 279  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
261 .. 279  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
297 .. 315  =  19 bp
bacterial operator O2 for the tetR and tetA genes
tet operator
297 .. 315  =  19 bp
bacterial operator O2 for the tetR and tetA genes
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