RNAi-mediated knockdown of MECP2. MECP2/MECP2 expression following RNAi-induced MeCP2 (e1 and e2) knockdown. Cells were harvested at time points over a 2-week period. (a) MECP2 mRNA expression in SH-SY5Y cells. Transcript levels were assessed by Real-Time qPCR and compared to the expression of GAPDH in each sample. MECP2 expression was quantitated at each time point, and presented as a percentage of the expression level at day 0, which was set at 100%. The knockdown sample (sh-MECP2) displays a 75-80% reduction in MECP2 expression when compared to the negative control (mut-sh MECP2) or to untreated cells (data not shown). MECP2 mRNA expression in the negative control remained above 95% greater than all time points; (b) MECP2 protein expression is reduced by 90% at Day 3. Each protein sample was divided in two and loaded onto duplicate gels, one immunoblotted with α-MECP2 and the other with α-actin.