Creating an miR30-based shRNA vector.

Generating expression constructs for artificial microRNAs (miRNAs) is relatively straightforward. This protocol describes the creation of miR-30-based short hairpin RNA (shRNA) cassettes that are compatible with a number of standard vector systems. The principles outlined here can also be easily applied to other miRNA scaffolds or to simple snapback shRNAs. It is important to note that one must understand the processing of the artificial scaffold and be able to predict precisely the small RNAs that will be generated. Otherwise, no design principles can be effectively applied and the probability that any individual shRNA clone will work effectively will be greatly reduced.