The pTet-t TS Vector, designed for use with the Tet-On TM Gene Expression System, prevent sun regulated gene expression in the absence of the inducing agent Doxycycline (Dox).

pTet-t TS encodes the tetracycline-controlled transcriptional silencer (tTS), which is a fusion of the tet repressor protein (TetR) and the KRAB-AB silencing domain of the Kid-1 protein (SDKid-1).

USE:
tTS will bind to the TRE in the presence of 0–10 ng/ml of Dox. It begins to dissociate from the TRE at concentrations >10 ng/ml. rtTA will bind to the TRE at >100 ng/ml of Dox. We recommend a working Dox concentration of 1 µg/ml for inducing gene expression.

For introduction during establishment of Tet-On cell lines
pTet-tTS does not contain a drug-resistance marker for clonal selection in mammalian cell culture. Therefore, we recommend co transfecting pet-tTS with the pTet- nVector, which contains the rtTA and the neomycin-resistance gene. Transfections should be performed at a 1:10 molar ratio (i.e., 1 µg pTet-On to 10 µg pTet-tTS).

Location of features:
Fragment containing PCMV promoter: 86–673
tTStranscriptional silencer:
Start codon: 775–777; stop codon: 1621–1623
Tet Repressor (TetR): 775–1407
SV40 nuclear localization sequence: 1408–1425
KRAB-AB Silencing Domain (SDKid-1): 1426–1623
Fragment containing SV40 poly-A sequence: 1647–2104
Col E1 origin of replication: 2454–3097
Ampicillin resistance (β-lactamase) gene:
Start codon: 4105–4103; stop codon: 3247–3245

Propagation in E. coli:
Suitable host strains: DH5α, HB101, and other general purpose strains.
Selectable marker: plasmid confers resistance to ampicillin (100 µg/ml) to E. coli hosts.
E. coli replication origin: Col E1
Copy number: low

PTET-TTS Vector