Real-Time qPCR as a Tool for Evaluating RNAi-Mediated Gene Silencing

To gain insight into the mechanism of action of MYCN and TP53 genes in neuroblastoma pathogenesis, and to create a model system for future exploration of targeted therapeutics, we decided to exploit RNAi as an experimental tool for suppressing the expression of MYCN and TP53. The MYCN gene was transiently suppressed using several 27-mer siLentMerTM* Dicer-substrate small interfering RNA (siRNA) duplexes, which are generally considered more effective at gene silencing than corresponding traditional 21-mer siRNAs (Kim et al. 2005). Stable knockdown of the TP53 gene was achieved using lentiviral-mediated short hairpin RNA (shRNA) interference. The CFX96TM real-time PCR detection system was used as a real-time detection platform.