Inducible Rassf5 and CreER expression

A mouse Rassf5 expression construct (Addgene) was cloned into tdTomato-C1 vector (Addgene) at EcoRI/SmaI sites. Rassf5-tdT was then cloned into a modified pLX Cre EF1 vector³ using the Zero Blunt TOPO kit (Invitrogen). To make inducible Rassf5 and CreER constructs, Rassf5-tdT was cloned into a pLKO2 TRE vector at NsiI/BspEI sites. CreER was cloned into a pLKO2 rtTA construct at SalI/NheI sites.RNA interference-mediated gene depletion was achieved using pLKO1 shRNA vectors from the mouse TRC1.0 shRNA library (Sigma-Aldrich). pLKO-Cre vectors were used to generate Cre-shRNA expression constructs (Beronja et al., 2013 (link); Beronja et al., 2010 (link)).