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Plasmid Files

ptTS-Neo

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

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ptTS-Neo Sequence and MapptTS-Neo.dna
Map and Sequence File   
Sequence Author:  Clontech
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 EcoNI (4) XmnI (5977) PvuI (5748) BsaI (5439) AhdI (5378) Bpu10I (4182) HindIII (4159) PflMI (3912) BglII (3705) Bsu36I (3635) ApaI (3535) PspOMI (3531) PsiI (3364) MfeI (3356) NsiI (61) SfiI (192) StuI (238) BsaBI * (276) EagI (323) KasI (416) NarI (417) SfoI (418) PluTI (420) PflFI - Tth111I (535) RsrII (933) BstBI (1099) BsrGI (1407) SpeI (1463) SnaBI (1804) Eco53kI (2030) SacI (2032) BsmBI (2053) BbsI (2098) SacII (2119) XbaI (2138) PmlI (2266) NheI (2785) BmtI (2789) AvaI - BsoBI (2829) BmeT110I (2830) BamHI (2971) KflI (3014) ptTS-Neo 6365 bp
EcoNI  (4)
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.
XmnI  (5977)
1 site
G A A N N N N T T C C T T N N N N A A G
PvuI  (5748)
1 site
C G A T C G G C T A G C
BsaI  (5439)
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  (5378)
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.
Bpu10I  (4182)
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.
HindIII  (4159)
1 site
A A G C T T T T C G A A
PflMI  (3912)
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.
BglII  (3705)
1 site
A G A T C T T C T A G A
Bsu36I  (3635)
1 site
C C T N A G G G G A N T C C

Sticky ends from different Bsu36I sites may not be compatible.
ApaI  (3535)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PspOMI  (3531)
1 site
G G G C C C C C C G G G
PsiI  (3364)
1 site
T T A T A A A A T A T T
MfeI  (3356)
1 site
C A A T T G G T T A A C
NsiI  (61)
1 site
A T G C A T T A C G T A
SfiI  (192)
1 site
G G C C N N N N N G G C C C C G G N N N N N C C G G

Efficient cleavage requires at least two copies of the SfiI
recognition sequence.
Sticky ends from different SfiI sites may not be compatible.
StuI  (238)
1 site
A G G C C T T C C G G A
BsaBI  (276)
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.
EagI  (323)
1 site
C G G C C G G C C G G C
KasI  (416)
1 site
G G C G C C C C G C G G
NarI  (417)
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  (418)
1 site
G G C G C C C C G C G G
PluTI  (420)
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.
PflFI  (535)
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  (535)
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.
RsrII  (933)
1 site
C G G W C C G G C C W G G C

Efficient cleavage requires at least two copies of the RsrII
recognition sequence.
Sticky ends from different RsrII sites may not be compatible.
For full activity, add fresh DTT.
BstBI  (1099)
1 site
T T C G A A A A G C T T
BsrGI  (1407)
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  (1463)
1 site
A C T A G T T G A T C A
SnaBI  (1804)
1 site
T A C G T A A T G C A T
Eco53kI  (2030)
1 site
G A G C T C C T C G A G
SacI  (2032)
1 site
G A G C T C C T C G A G
BsmBI  (2053)
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  (2098)
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.
SacII  (2119)
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.
XbaI  (2138)
1 site
T C T A G A A G A T C T
PmlI  (2266)
1 site
C A C G T G G T G C A C

PmlI gradually loses activity when stored at -20°C.
NheI  (2785)
1 site
G C T A G C C G A T C G
BmtI  (2789)
1 site
G C T A G C C G A T C G
AvaI  (2829)
1 site
C Y C G R G G R G C Y C

Sticky ends from different AvaI sites may not be compatible.
BsoBI  (2829)
1 site
C Y C G R G G R G C Y C

Sticky ends from different BsoBI sites may not be compatible.
BsoBI is typically used at 37°C, but can be used at temperatures
up to 65°C.
BmeT110I  (2830)
1 site
C Y C G R G G R G C Y C
BamHI  (2971)
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.
KflI  (3014)
1 site
G G G W C C C C C C W G G G

Sticky ends from different KflI sites may not be compatible.
AmpR
5305 .. 6165  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 2:  
   5305 .. 6096  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
5305 .. 6165  =  861 bp
286 amino acids  =  31.5 kDa
   Segment 1:  signal sequence  
   6097 .. 6165  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
AmpR
5305 .. 6165  =  861 bp
286 amino acids  =  31.5 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and
related antibiotics
tTS
2135 .. 2983  =  849 bp
282 amino acids  =  31.9 kDa
   Segment 1:  TetR  
   2135 .. 2767  =  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
2135 .. 2983  =  849 bp
282 amino acids  =  31.9 kDa
   Segment 2:  NLS  
   2768 .. 2788  =  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
2135 .. 2983  =  849 bp
282 amino acids  =  31.9 kDa
   Segment 3:  KRAB silencing domain  
   2789 .. 2983  =  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
2135 .. 2983  =  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
NeoR/KanR
289 .. 1083  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin)
NeoR/KanR
289 .. 1083  =  795 bp
264 amino acids  =  29.0 kDa
Product: aminoglycoside phosphotransferase from
Tn5
confers resistance to neomycin, kanamycin, and
G418 (Geneticin)
ori
4546 .. 5134  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
ori
4546 .. 5134  =  589 bp
high-copy-number ColE1/pMB1/pBR322/pUC origin
of replication
β-globin intron
3004 .. 3576  =  573 bp
intron from rabbit β-globin gene
β-globin intron
3004 .. 3576  =  573 bp
intron from rabbit β-globin gene
CMV enhancer
1449 .. 1828  =  380 bp
human cytomegalovirus immediate early enhancer
CMV enhancer
1449 .. 1828  =  380 bp
human cytomegalovirus immediate early enhancer
CMV promoter
1829 .. 2032  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
CMV promoter
1829 .. 2032  =  204 bp
human cytomegalovirus (CMV) immediate early
promoter
SV40 promoter
58 .. 254  =  197 bp
SV40 early promoter
SV40 promoter
58 .. 254  =  197 bp
SV40 early promoter
AmpR promoter
6166 .. 6270  =  105 bp
AmpR promoter
6166 .. 6270  =  105 bp
β-globin poly(A) signal
3773 .. 3828  =  56 bp
rabbit β-globin polyadenylation signal
β-globin poly(A) signal
3773 .. 3828  =  56 bp
rabbit β-globin polyadenylation signal
HSV TK poly(A) signal
1315 .. 1363  =  49 bp
herpesvirus thymidine kinase polyadenylation signal
HSV TK poly(A) signal
1315 .. 1363  =  49 bp
herpesvirus thymidine kinase polyadenylation signal
SV40 ori
105 .. 240  =  136 bp
SV40 origin of replication
SV40 ori
105 .. 240  =  136 bp
SV40 origin of replication
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