pTRE-Dual2

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

Sequence Author: Clontech (TaKaRa)

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ZraI (3813) SspI (3697) EarI (3688) ScaI (3373) PvuI (3263) FspI (3115) AseI (3065) BsaI (2954) PsiI (1817) AatII (3815) Eco53kI (292) SacI (294) EcoRI (323) PstI - SbfI (690) BbvCI - Bpu10I (873) PvuII (913) BsgI (929) SgrAI (1016) XcmI (1021) BsrGI (1035) NheI (1046) BmtI (1050) PspOMI (1166) ApaI (1170) AvrII (1204) HindIII (1277) PmlI (1369) BfuAI - BspMI - PaqCI (1392) DraIII (1416) Acc65I (1494) KpnI (1498) BmgBI (1596) EagI - NotI - SacII (1641) BglII (1649) BamHI (1655) BspDI - ClaI (1662) SalI (1667) AccI (1668) EcoRV (1675) NdeI (1680) XbaI (1685) MfeI (1784) HpaI (1797) pTRE-Dual2 3884 bp
ZraI  (3813)
1 site
G A C G T C C T G C A G
SspI  (3697)
1 site
A A T A T T T T A T A A
EarI  (3688)
1 site
C T C T T C N G A G A A G N N N N

Cleavage may be enhanced when more than one copy of the EarI recognition sequence is present.
Sticky ends from different EarI sites may not be compatible.
ScaI  (3373)
1 site
A G T A C T T C A T G A
PvuI  (3263)
1 site
C G A T C G G C T A G C
FspI  (3115)
1 site
T G C G C A A C G C G T
AseI  (3065)
1 site
A T T A A T T A A T T A
BsaI  (2954)
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.
PsiI  (1817)
1 site
T T A T A A A A T A T T
AatII  (3815)
1 site
G A C G T C C T G C A G
Eco53kI  (292)
1 site
G A G C T C C T C G A G
SacI  (294)
1 site
G A G C T C C T C G A G
EcoRI  (323)
1 site
G A A T T C C T T A A G
PstI  (690)
1 site
C T G C A G G A C G T C
SbfI  (690)
1 site
C C T G C A G G G G A C G T C C
BbvCI  (873)
1 site
C C T C A G C G G A G T C G
Bpu10I  (873)
1 site
C C T N A G C G G A N T C G

Cleavage may be enhanced when more than one copy of the Bpu10I recognition sequence is present.
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.
PvuII  (913)
1 site
C A G C T G G T C G A C
BsgI  (929)
1 site
G T G C A G ( N ) 14 N N C A C G T C ( N ) 14

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).
SgrAI  (1016)
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.
XcmI  (1021)
1 site
C C A N N N N N N N N N T G G G G T N N N N N N N N N A C C

The 1-base overhangs produced by XcmI may be hard to ligate.
Sticky ends from different XcmI sites may not be compatible.
BsrGI  (1035)
1 site
T G T A C A A C A T G T

BsrGI is typically used at 37°C, but is even more active at 60°C.
NheI  (1046)
1 site
G C T A G C C G A T C G
BmtI  (1050)
1 site
G C T A G C C G A T C G
PspOMI  (1166)
1 site
G G G C C C C C C G G G
ApaI  (1170)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
AvrII  (1204)
1 site
C C T A G G G G A T C C
HindIII  (1277)
1 site
A A G C T T T T C G A A
PmlI  (1369)
1 site
C A C G T G G T G C A C
BfuAI  (1392)
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  (1392)
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.
PaqCI  (1392)
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 PaqCI recognition sequence.
Sticky ends from different PaqCI sites may not be compatible.
Cleavage can be improved with PaqCI Activator.
DraIII  (1416)
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.
Acc65I  (1494)
1 site
G G T A C C C C A T G G
KpnI  (1498)
1 site
G G T A C C C C A T G G
BmgBI  (1596)
1 site
C A C G T C G T G C A G

This recognition sequence is asymmetric, so ligating blunt ends generated by BmgBI will not always regenerate a BmgBI site.
EagI  (1641)
1 site
C G G C C G G C C G G C
NotI  (1641)
1 site
G C G G C C G C C G C C G G C G
SacII  (1641)
1 site
C C G C G G G G C G C C

Efficient cleavage requires at least two copies of the SacII recognition sequence.
BglII  (1649)
1 site
A G A T C T T C T A G A
BamHI  (1655)
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.
BspDI  (1662)
1 site
A T C G A T T A G C T A
ClaI  (1662)
1 site
A T C G A T T A G C T A
SalI  (1667)
1 site
G T C G A C C A G C T G
AccI  (1668)
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  (1675)
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.
NdeI  (1680)
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  (1685)
1 site
T C T A G A A G A T C T
MfeI  (1784)
1 site
C A A T T G G T T A A C
HpaI  (1797)
1 site
G T T A A C C A A T T G
AmpR
2820 .. 3680  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
   Segment 2:  
   2820 .. 3611  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2820 .. 3680  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
   Segment 1:  signal sequence  
   3612 .. 3680  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
2820 .. 3680  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
mCherry
335 .. 1045  =  711 bp
236 amino acids  =  26.7 kDa
Product: monomeric derivative of DsRed fluorescent protein
mammalian codon-optimized
mCherry
335 .. 1045  =  711 bp
236 amino acids  =  26.7 kDa
Product: monomeric derivative of DsRed fluorescent protein
mammalian codon-optimized
ori
2061 .. 2649  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
ori
2061 .. 2649  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin of replication
IRES2
1053 .. 1639  =  587 bp
3 segments
   Segment 1:  
   1053 .. 1627  =  575 bp
internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)
IRES2
1053 .. 1639  =  587 bp
3 segments
   Segment 2:  ATG  
   1628 .. 1630  =  3 bp
internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)
IRES2
1053 .. 1639  =  587 bp
3 segments
   Segment 3:  
   1631 .. 1639  =  9 bp
internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)
IRES2
1053 .. 1639  =  587 bp
3 segments
internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)
tight TRE promoter
4 .. 318  =  315 bp
Tet-responsive promoter PTight, consisting of seven tet operator sequences followed by the minimal CMV promoter
tight TRE promoter
4 .. 318  =  315 bp
Tet-responsive promoter PTight, consisting of seven tet operator sequences followed by the minimal CMV promoter
AmpR promoter
3681 .. 3785  =  105 bp
AmpR promoter
3681 .. 3785  =  105 bp
SV40 poly(A) signal
1798 .. 1879  =  82 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1798 .. 1879  =  82 bp
SV40 polyadenylation signal
MCS
1640 .. 1690  =  51 bp
multiple cloning site
MCS
1640 .. 1690  =  51 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
ATG
1628 .. 1630  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
1628 .. 1630  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ORF:  70 .. 360  =  291 bp
ORF:  96 amino acids  =  11.1 kDa
ORF:  2950 .. 3216  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  3835 .. 202  =  252 bp
ORF:  83 amino acids  =  9.8 kDa
ORF:  335 .. 1045  =  711 bp
ORF:  236 amino acids  =  26.7 kDa
ORF:  1536 .. 1775  =  240 bp
ORF:  79 amino acids  =  9.1 kDa
ORF:  2820 .. 3680  =  861 bp
ORF:  286 amino acids  =  31.5 kDa
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Download pTRE-Dual2.dna file

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