Post-transcriptional gene silencing is a powerful tool to reveal gene function. In this chapter, we have used gene-array technology to analyze changes in gene expression after specific targeting of the protein kinase Calpha isoform with small interfering RNAs (siRNAs). Expression profiles were investigated on a human gene array of 588 key genes involved in DNA synthesis, apoptosis, cell-cell communication, intracellular signal-transduction pathways, cell-surface molecules and transcription factors. The data indicate that siRNAs are an ideal tool for target validation, and show that RNA-dependent RNA polymerase is not required for RNAi in mammalian cells.