Figure 5. Conditional silencing using the selectable one-vector GLTR-X system.

U2OS cells were infected with pGLTR-X-Puro-CDC27. Forty eight hours after infection cells were selected for puromycin resistance (1 µg/ml). Resistant cells were cultured in the presence or absence of 1 µg/ml Dox for 3 days and analysed by phase contrast microscopy (A). For time-course experiments (B), cells were treated with 1 µg/ml Dox and samples taken at the indicated time points for immunoblotting for CDC27 levels as well as GAPDH as loading control. For dose-response analysis (C), cells were treated with increasing doses of Dox for 3 days and total cell extracts analysed for CDC27 and GAPDH expression.