On targetplus non targeting pool d 001810

siRNA oligonucleotide sequences were used to target p53, p63, p73 and Puma (Dharmacon, ONTARGETplus SMARTpool). For negative control, a nontarget sequence was used (Dharmacon, ON-TARGETplus Non-Targeting Pool, D-001810). Cortical neurons dissociated cells were transfected with siRNA 3 days after plating, using the protocol supplied with DharmaFECT 4 (Dharmacon, T-2004–03) and performed as previously described before (Maor-Nof et al., 2013 (link)) with minor modifications. Briefly, siRNA and the transfection reagent were each diluted separately into NB medium without serum and antibiotics for 5 min; then, the siRNA was added to the medium with the transfection reagent. After an additional 20 min incubation, the transfection reagent siRNA complex was added to the dissociated cells and grown in NB medium without serum and antibiotics. 16h later, the transfection reagent was removed by replacing the medium with a complete medium and the neurons were cultured for an additional 48 hr. Lentivirus transduction was performed after. The final concentration of the siRNA was 0.1 mM. The level of the target protein Puma, reduced by its specific siRNA treatment, was evaluated by western blot analysis.