Project description:Basal gene expression of T47D-MTVL human breast cancer cells growing in normal conditions Breast cancer cells T47D-MTVL cells were grown in rich media and total RNA was extracted to monitor basal gene expression
Project description:H1.2 (and control) knock-down cell lines were established from T47D-MTVL cells: Human breast cancer cell line T47D modified to contain one stably integrated copy of Luciferase reporter gene driven by the Mouse Mammary Tumor Virus promoter, named T47D-MTVL (Truss, M., J. Bartsch, A. Schelbert, R. J. Hache, and M. Beato. 1995). Initially, a cell line expressing the Dox-responsive KRAB repressor and RedFP (ptTR-KRAB-Red) was generated. Then, this cell line was infected with viruses for expression of the different H1 variants shRNAs (pLVTHM). The inducible knocked-down cell lines were sorted in a FACSvantageSE (Becton Dickinson) for RedFP-positive and GFP-positive fluorescence after 3 days of Dox treatment. Then cells were amplified in the absence of Dox until an experiment was performed. Plasmids for the lentivirus vector-mediated drug-inducible RNA interference system (pLVTHM, ptTR-KRAB-Red, pCMC-R8.91 and pMD.G) were provided by D. Trono (University of Geneva) (Wiznerowicz M & Trono D (2003) Conditional suppression of cellular genes: lentivirus vector-mediated drug-inducible RNA interference. J Virol 77: 8957-8961). Keywords: Gene expression - T47D-MTVL-control sh/sh H1.2 +/- Dox induction
