Effective target silencing and oncogene expression can be obtained from one unique mRNA. (A) Retroviral vectors used to deliver shRNAs to mammalian cells. PLMP vector encodes for a miR30-based target-specific shRNA under LTR promoter control. PMmiR^TOI–BCR-ABL vector encodes for target-specific miR and p185 BCR-ABL. Provirus layouts are shown with open arrows indicating active promoters and 2 inverted block arrows representing shRNA stem sequence. (B) Western blot analysis of NIH/3T3 cells transduced with different Raf1 miR in pLMP after puromycin selection. (C) Ba/F3 cells retrovirally infected with pMmiR^Raf1–BCR-ABL or pMmiR^Ctrl–BCR-ABL construct. (D) Western blot analysis of NIH/3T3 cells transduced with p53 miR in pLMP after puromycin selection. To induce p53, cells were UV irradiated 4 hours prior harvesting. (E) Western blot analysis of Ba/F3 cells retrovirally infected with pMmiR^p53–BCR-ABL or pMmiR^Ctrl–BCR-ABL. (F-G) Raf1 knockdown disrupts BCR-ABL–dependent MAPK signaling. 32D (F) or Ba/F3 cells (G) retrovirally infected with pMmiR^Ctrl–BCR-ABL or pMmiR^Raf1–BCR-ABL were maintained in IL-3–containing medium, then starved for 4 hours and analyzed for MAPK signaling by Western blot as indicated.