KO_FG12_R2
MTECs were isolated, cultured, and differentiated as previously described (Lam et al., 2011; Vladar and Brody, 2013). MTECs harvested from p73+/+ and p73-/- mice were infected twice (at time of plating and 12 h later) with TAp73b and Fg12-CMV control expression vectors as described previously (Rosenbluth et al., 2011; Rosenbluth et al., 2008). MTECs were grown as submerged cultures on collagen-coated plates for 3 d in MTEC growth media (DMEM/F12 supplemented with HEPES, L-glutamine, NaHCO3, penicillin-streptomycin, fungizone, insulin, transferrin, cholera toxin, EGF, bovine pituitary extract and FBS). As well as MTECs were grown as submerged cultures on collagen-coated plates for 3 d in DMEM and then switched to differentiation media containing 2% NuSerum for 3 d (Lam et al., 2011; Vladar and Brody, 2013). MTECs were grown in transwells containing MTEC growth media for 5 days and then lifted to air-liquid interface for 24 h in differentiation media followed by RNA harvest using the Aurum Total RNA Mini kit (Bio-Rad). Duplicate samples for each of the four genotype/expression conditions (n=8 total) were submitted to the HudsonAlpha Genomic Services Lab for library preparation and RNA-seq.