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Plasmid Files

pTet-tTS

Vector for expressing tTS, which represses expression from Tet-responsive promoters in the absence of doxycycline.

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pTet-tTS Sequence and MappTet-tTS.dna
Map and Sequence File   
Sequence Author:  Clontech
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 PaeR7I - PspXI - XhoI (5) XmnI (3931) ScaI (3812) PvuI (3702) BsaI (3393) AhdI (3332) AlwNI (2855) PspFI (2747) BseYI (2743) DrdI (2547) AflIII - PciI (2439) BspQI - SapI (2323) PvuII (2263) Bpu10I (2136) BsrGI (51) SpeI (107) NdeI (342) SnaBI (448) Eco53kI (674) SacI (676) BsmBI (697) SacII (763) EcoRI (770) XbaI (782) PmlI (910) PfoI * (1014) BstXI (1061) NheI (1429) BmtI (1433) BspDI - ClaI (1629) PsiI (1760) HpaI (1780) MfeI (1789) BsaBI * (1881) KasI (1994) NarI (1995) SfoI (1996) PluTI (1998) SexAI * (2011) BstEII (2026) NgoMIV (2070) NaeI (2072) HindIII (2113) pTet-tTS 4319 bp
PaeR7I  (5)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (5)
1 site
V C T C G A G B B G A G C T C V
XhoI  (5)
1 site
C T C G A G G A G C T C
XmnI  (3931)
1 site
G A A N N N N T T C C T T N N N N A A G
ScaI  (3812)
1 site
A G T A C T T C A T G A
PvuI  (3702)
1 site
C G A T C G G C T A G C
BsaI  (3393)
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  (3332)
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.
AlwNI  (2855)
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.
PspFI  (2747)
1 site
C C C A G C G G G T C G
BseYI  (2743)
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.
DrdI  (2547)
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.
AflIII  (2439)
1 site
A C R Y G T T G Y R C A

Sticky ends from different AflIII sites may not be compatible.
PciI  (2439)
1 site
A C A T G T T G T A C A

PciI is inhibited by nonionic detergents.
BspQI  (2323)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different BspQI sites may not be compatible.
SapI  (2323)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different SapI sites may not be compatible.
SapI gradually settles in solution, so a tube of SapI should be
mixed before removing an aliquot.
PvuII  (2263)
1 site
C A G C T G G T C G A C
Bpu10I  (2136)
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  (51)
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.
SpeI  (107)
1 site
A C T A G T T G A T C A
NdeI  (342)
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.
SnaBI  (448)
1 site
T A C G T A A T G C A T
Eco53kI  (674)
1 site
G A G C T C C T C G A G
SacI  (676)
1 site
G A G C T C C T C G A G
BsmBI  (697)
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  (763)
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.
EcoRI  (770)
1 site
G A A T T C C T T A A G
XbaI  (782)
1 site
T C T A G A A G A T C T
PmlI  (910)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
PfoI  (1014)
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.
BstXI  (1061)
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.
NheI  (1429)
1 site
G C T A G C C G A T C G
BmtI  (1433)
1 site
G C T A G C C G A T C G
BspDI  (1629)
1 site
A T C G A T T A G C T A
ClaI  (1629)
1 site
A T C G A T T A G C T A
PsiI  (1760)
1 site
T T A T A A A A T A T T
HpaI  (1780)
1 site
G T T A A C C A A T T G
MfeI  (1789)
1 site
C A A T T G G T T A A C
BsaBI  (1881)
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.
KasI  (1994)
1 site
G G C G C C C C G C G G
NarI  (1995)
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  (1996)
1 site
G G C G C C C C G C G G
PluTI  (1998)
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.
SexAI  (2011)
1 site
A C C W G G T T G G W C C A
* Blocked by Dcm methylation.
Sticky ends from different SexAI sites may not be compatible.
BstEII  (2026)
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.
NgoMIV  (2070)
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.
NaeI  (2072)
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.
HindIII  (2113)
1 site
A A G C T T T T C G A A
AmpR
3259 .. 4119  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 2:  
   3259 .. 4050  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3259 .. 4119  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 1:  signal sequence  
   4051 .. 4119  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
3259 .. 4119  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
tTS
779 .. 1627  =  849 bp
282 amino acids  =  31.9 kDa
   Segment 1:  TetR  
   779 .. 1411  =  633 bp
   211 amino acids  =  23.6 kDa
Product: fusion of a hybrid tetracycline repressor
TetR with a nuclear localization signal and a KRAB
silencing domain
silences Tet-responsive promoters in the absence
but not the presence of doxycycline
tTS
779 .. 1627  =  849 bp
282 amino acids  =  31.9 kDa
   Segment 2:  NLS  
   1412 .. 1432  =  21 bp
   7 amino acids  =  897.2 Da
Product: fusion of a hybrid tetracycline repressor
TetR with a nuclear localization signal and a KRAB
silencing domain
silences Tet-responsive promoters in the absence
but not the presence of doxycycline
tTS
779 .. 1627  =  849 bp
282 amino acids  =  31.9 kDa
   Segment 3:  KRAB silencing domain  
   1433 .. 1627  =  195 bp
   64 amino acids  =  7.4 kDa
Product: fusion of a hybrid tetracycline repressor
TetR with a nuclear localization signal and a KRAB
silencing domain
silences Tet-responsive promoters in the absence
but not the presence of doxycycline
tTS
779 .. 1627  =  849 bp
282 amino acids  =  31.9 kDa
3 segments
Product: fusion of a hybrid tetracycline repressor
TetR with a nuclear localization signal and a KRAB
silencing domain
silences Tet-responsive promoters in the absence
but not the presence of doxycycline
ori
2500 .. 3088  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
2500 .. 3088  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
CMV enhancer
93 .. 472  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
93 .. 472  =  380 bp
human cytomegalovirus immediate early enhancer
CMV promoter
473 .. 676  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
473 .. 676  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
SV40 poly(A) signal
1646 .. 1780  =  135 bp
SV40 polyadenylation signal
SV40 poly(A) signal
1646 .. 1780  =  135 bp
SV40 polyadenylation signal
AmpR promoter
4120 .. 4224  =  105 bp
AmpR promoter
4120 .. 4224  =  105 bp
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