pTet-DualOFF

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

Sequence Author: Clontech (TaKaRa)

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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 - Esp3I (538) SacII (604) BamHI (607) PvuII (621) MluI (625) StuI (704) NruI (914) FseI (1258) TspMI - XmaI (1372) SmaI (1374) EcoRI (1384) AvrII (1543) PaqCI (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-DualOFF 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

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  (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

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.
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.
BsmBI-v2 is an improved version of BsmBI.
Esp3I  (538)
1 site
C G T C T C N G C A G A G N ( N ) 4

Sticky ends from different Esp3I 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

Cleavage may be enhanced when more than one copy of the XmaI recognition sequence is present.
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
PaqCI  (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 PaqCI recognition sequence.
Sticky ends from different PaqCI sites may not be compatible.
Cleavage can be improved with PaqCI Activator.
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
2 segments
   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
2 segments
   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
tTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
8 segments
   Segment 1:  TetR  
   634 .. 1251  =  618 bp
   206 amino acids  =  23.3 kDa
Product: improved tetracycline-controlled transactivator
In the Tet-Off® system, tTA binds to the tetracycline response element and stimulates transcription only in the absence of tetracycline or doxycycline.
tTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
8 segments
   Segment 2:  
   1252 .. 1257  =  6 bp
   2 amino acids  =  172.2 Da
Product: improved tetracycline-controlled transactivator
In the Tet-Off® system, tTA binds to the tetracycline response element and stimulates transcription only in the absence of tetracycline or doxycycline.
tTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
8 segments
   Segment 3:  VP16 minimal transcriptional activation domain  
   1258 .. 1293  =  36 bp
   12 amino acids  =  1.4 kDa
Product: improved tetracycline-controlled transactivator
In the Tet-Off® system, tTA binds to the tetracycline response element and stimulates transcription only in the absence of tetracycline or doxycycline.
tTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
8 segments
   Segment 4:  
   1294 .. 1296  =  3 bp
   1 amino acid  =  115.1 Da
Product: improved tetracycline-controlled transactivator
In the Tet-Off® system, tTA binds to the tetracycline response element and stimulates transcription only in the absence of tetracycline or doxycycline.
tTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
8 segments
   Segment 5:  VP16 minimal transcriptional activation domain  
   1297 .. 1332  =  36 bp
   12 amino acids  =  1.4 kDa
Product: improved tetracycline-controlled transactivator
In the Tet-Off® system, tTA binds to the tetracycline response element and stimulates transcription only in the absence of tetracycline or doxycycline.
tTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
8 segments
   Segment 6:  
   1333 .. 1335  =  3 bp
   1 amino acid  =  115.1 Da
Product: improved tetracycline-controlled transactivator
In the Tet-Off® system, tTA binds to the tetracycline response element and stimulates transcription only in the absence of tetracycline or doxycycline.
tTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
8 segments
   Segment 7:  VP16 minimal transcriptional activation domain  
   1336 .. 1371  =  36 bp
   12 amino acids  =  1.3 kDa
Product: improved tetracycline-controlled transactivator
In the Tet-Off® system, tTA binds to the tetracycline response element and stimulates transcription only in the absence of tetracycline or doxycycline.
tTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
8 segments
   Segment 8:  
   1372 .. 1380  =  9 bp
   2 amino acids  =  172.2 Da
Product: improved tetracycline-controlled transactivator
In the Tet-Off® system, tTA binds to the tetracycline response element and stimulates transcription only in the absence of tetracycline or doxycycline.
tTA-Advanced
634 .. 1380  =  747 bp
248 amino acids  =  27.7 kDa
8 segments
Product: improved tetracycline-controlled transactivator
In the Tet-Off® system, tTA binds to the tetracycline response element and stimulates transcription only in the absence of tetracycline or doxycycline.
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
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
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
IRES2
1392 .. 1978  =  587 bp
3 segments
   Segment 1:  
   1392 .. 1966  =  575 bp
internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)
IRES2
1392 .. 1978  =  587 bp
3 segments
   Segment 2:  ATG  
   1967 .. 1969  =  3 bp
internal ribosome entry site (IRES) of the encephalomyocarditis virus (EMCV)
IRES2
1392 .. 1978  =  587 bp
3 segments
   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)
ORF:  634 .. 1380  =  747 bp
ORF:  248 amino acids  =  27.7 kDa
ORF:  791 .. 1093  =  303 bp
ORF:  100 amino acids  =  11.2 kDa
ORF:  1967 .. 2674  =  708 bp
ORF:  235 amino acids  =  26.5 kDa
ORF:  3972 .. 4238  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  3842 .. 4702  =  861 bp
ORF:  286 amino acids  =  31.5 kDa
ORF:  907 .. 1170  =  264 bp
ORF:  87 amino acids  =  9.8 kDa
ORF:  2356 .. 2649  =  294 bp
ORF:  97 amino acids  =  10.9 kDa
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Download pTet-DualOFF.dna file

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Individual Sequences & Maps

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