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Plasmid Files

pTet-DualON

Vector for co-expressing ZsGreen1 with the Tet-On® Advanced transcriptional activator.

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pTet-DualON Sequence and MappTet-DualON.dna
Map and Sequence File   
Sequence Author:  Clontech
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 SspI (4719) EarI (4710) ScaI (4395) PvuI (4285) FspI (4137) AseI (4087) BsaI (3976) AlwNI (3438) DrdI (3130) PsiI (2839) HpaI (2819) BsaBI * (2718) EcoRV (2703) BspDI - ClaI (2684) NotI (2676) Bpu10I (2671) BsrGI (2529) PaeR7I - PspXI - XhoI (1) SnaBI (289) Eco53kI (515) SacI (517) BsmBI (538) SacII (604) BamHI (607) PvuII (621) MluI (625) StuI (704) NruI (914) FseI (1258) TspMI - XmaI (1372) SmaI (1374) EcoRI (1384) AvrII (1543) AarI (1731) DraIII (1755) Acc65I (1833) KpnI (1837) PflMI (1845) MscI (1970) BstXI (1978) BclI * (2061) SgrAI (2066) PflFI - Tth111I (2213) KasI (2282) NarI (2283) SfoI (2284) PluTI (2286) pTet-DualON 4906 bp
SspI  (4719)
1 site
A A T A T T T T A T A A
EarI  (4710)
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.
ScaI  (4395)
1 site
A G T A C T T C A T G A
PvuI  (4285)
1 site
C G A T C G G C T A G C
FspI  (4137)
1 site
T G C G C A A C G C G T
AseI  (4087)
1 site
A T T A A T T A A T T A
BsaI  (3976)
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.
AlwNI  (3438)
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.
DrdI  (3130)
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.
PsiI  (2839)
1 site
T T A T A A A A T A T T
HpaI  (2819)
1 site
G T T A A C C A A T T G
BsaBI  (2718)
1 site
G A T N N N N A T C C T A N N N N T A G
* Blocked by Dam methylation.
EcoRV  (2703)
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.
BspDI  (2684)
1 site
A T C G A T T A G C T A
ClaI  (2684)
1 site
A T C G A T T A G C T A
NotI  (2676)
1 site
G C G G C C G C C G C C G G C G
Bpu10I  (2671)
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.
BsrGI  (2529)
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.
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
SnaBI  (289)
1 site
T A C G T A A T G C A T
Eco53kI  (515)
1 site
G A G C T C C T C G A G
SacI  (517)
1 site
G A G C T C C T C G A G
BsmBI  (538)
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.
SacII  (604)
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.
BamHI  (607)
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.
PvuII  (621)
1 site
C A G C T G G T C G A C
MluI  (625)
1 site
A C G C G T T G C G C A
StuI  (704)
1 site
A G G C C T T C C G G A
NruI  (914)
1 site
T C G C G A A G C G C T
FseI  (1258)
1 site
G G C C G G C C C C G G C C G G

FseI gradually loses activity when stored at -20°C.
TspMI  (1372)
1 site
C C C G G G G G G C C C
XmaI  (1372)
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  (1374)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
EcoRI  (1384)
1 site
G A A T T C C T T A A G
AvrII  (1543)
1 site
C C T A G G G G A T C C
AarI  (1731)
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.
DraIII  (1755)
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  (1833)
1 site
G G T A C C C C A T G G
KpnI  (1837)
1 site
G G T A C C C C A T G G
PflMI  (1845)
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.
MscI  (1970)
1 site
T G G C C A A C C G G T
BstXI  (1978)
1 site
C C A N N N N N N T G G G G T N N N N N N A C C

Sticky ends from different BstXI sites may not be compatible.
BclI  (2061)
1 site
T G A T C A A C T A G T
* Blocked by Dam methylation.
BclI is typically used at 50-55°C, but is 50% active at 37°C.
SgrAI  (2066)
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.
PflFI  (2213)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by PflFI may be hard to ligate.
Sticky ends from different PflFI sites may not be compatible.
Tth111I  (2213)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by Tth111I may be hard to ligate.
Sticky ends from different Tth111I sites may not be compatible.
KasI  (2282)
1 site
G G C G C C C C G C G G
NarI  (2283)
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  (2284)
1 site
G G C G C C C C G C G G
PluTI  (2286)
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.
AmpR
3842 .. 4702  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 2:  
   3842 .. 4633  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3842 .. 4702  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 1:  signal sequence  
   4634 .. 4702  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3842 .. 4702  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
rtTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
   Segment 1:  rTetR  
   634 .. 1251  =  618 bp
   206 amino acids  =  23.2 kDa
Product: improved tetracycline-controlled
transactivator
binds to a Tet-responsive element in the presence
but not the absence of doxycycline
rtTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
   Segment 2:  
   1252 .. 1257  =  6 bp
   2 amino acids  =  172.2 Da
Product: improved tetracycline-controlled
transactivator
binds to a Tet-responsive element in the presence
but not the absence of doxycycline
rtTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
   Segment 3:  VP16 minimal transcriptional activation domain  
   1258 .. 1293  =  36 bp
   12 amino acids  =  1.4 kDa
Product: improved tetracycline-controlled
transactivator
binds to a Tet-responsive element in the presence
but not the absence of doxycycline
rtTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
   Segment 4:  
   1294 .. 1296  =  3 bp
   1 amino acid  =  115.1 Da
Product: improved tetracycline-controlled
transactivator
binds to a Tet-responsive element in the presence
but not the absence of doxycycline
rtTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
   Segment 5:  VP16 minimal transcriptional activation domain  
   1297 .. 1332  =  36 bp
   12 amino acids  =  1.4 kDa
Product: improved tetracycline-controlled
transactivator
binds to a Tet-responsive element in the presence
but not the absence of doxycycline
rtTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
   Segment 6:  
   1333 .. 1335  =  3 bp
   1 amino acid  =  115.1 Da
Product: improved tetracycline-controlled
transactivator
binds to a Tet-responsive element in the presence
but not the absence of doxycycline
rtTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
   Segment 7:  VP16 minimal transcriptional activation domain  
   1336 .. 1371  =  36 bp
   12 amino acids  =  1.4 kDa
Product: improved tetracycline-controlled
transactivator
binds to a Tet-responsive element in the presence
but not the absence of doxycycline
rtTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
   Segment 8:  
   1372 .. 1380  =  9 bp
   2 amino acids  =  172.2 Da
Product: improved tetracycline-controlled
transactivator
binds to a Tet-responsive element in the presence
but not the absence of doxycycline
rtTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
8 segments
Product: improved tetracycline-controlled
transactivator
binds to a Tet-responsive element in the presence
but not the absence of doxycycline
ZsGreen1
1979 .. 2674  =  696 bp
231 amino acids  =  26.1 kDa
Product: Zoanthus green fluorescent protein
mammalian codon-optimized
ZsGreen1
1979 .. 2674  =  696 bp
231 amino acids  =  26.1 kDa
Product: Zoanthus green fluorescent protein
mammalian codon-optimized
ori
3083 .. 3671  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
3083 .. 3671  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
IRES2
1392 .. 1978  =  587 bp
   Segment 1:  
   1392 .. 1966  =  575 bp
internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)
IRES2
1392 .. 1978  =  587 bp
   Segment 2:  ATG  
   1967 .. 1969  =  3 bp
internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)
IRES2
1392 .. 1978  =  587 bp
   Segment 3:  
   1970 .. 1978  =  9 bp
internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)
IRES2
1392 .. 1978  =  587 bp
3 segments
internal ribosome entry site (IRES) of the
encephalomyocarditis virus (EMCV)
CMV enhancer
10 .. 313  =  304 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
10 .. 313  =  304 bp
human cytomegalovirus immediate early enhancer
CMV promoter
314 .. 517  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
314 .. 517  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
AmpR promoter
4703 .. 4807  =  105 bp
AmpR promoter
4703 .. 4807  =  105 bp
SV40 poly(A) signal
2820 .. 2901  =  82 bp
SV40 polyadenylation signal
SV40 poly(A) signal
2820 .. 2901  =  82 bp
SV40 polyadenylation signal
ATG
1967 .. 1969  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
ATG
1967 .. 1969  =  3 bp
1 amino acid  =  149.2 Da
Product: start codon
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