What is the best time-frame for monitoring siRNA-dependent decreases in target mRNA expression levels? In target protein levels?
Generally, target mRNA levels are decreased after 24 hours post-transfection following transfection of a gene-specific siRNA duplex. However, maximal silencing may be reached at a later time point, so it is advisable to assay target mRNA in a time-course study. In most cases, silencing will be maximal at 24 to 48 hours following transfection. Cellular target protein levels may persist after the mRNA has been silenced based on the stability of the protein in your experimental system. Protein levels should be examined starting at 24 hours and assayed until a minimum level is noted, often 48 to 96 hours or greater. As always, it is important to verify transfection efficiency using an appropriate positive control.