Expression of shRNA using intron splicing

RNA is considered a highly promising candidate for several applications such as gene knock-down, gene repair and gene therapy, where double-stranded RNA and RNA with catalytic activity are key players. A group I intron, a ribozyme catalyzing its own splicing reactions in the absence of any proteins, has generated interest for its potential utility in gene repair using trans-splicing. On the other hand, the induction of small interfering RNA, via double-stranded RNA cleavage in short hairpin RNA (shRNA) by the RNase* *enzyme DICER is a convenient and powerful mechanism for gene silencing. We constructed shRNA expression vectors directed against Firefly luciferase, in which the loop region of the shRNA was interrupted by an intron. The decreased levels of luciferase activity were measured in cultured cells as an index of the ribozyme splicing activity.