Ribozyme Expression Systems

We have succeeded in constructing an effective system for the expression of ribozymes under the control of a human tRNA^Val promoter, which ensures a high level of production of ribozymes in vivo. The engineered tRNA^Val -driven ribozymes, based on computer-predicted secondary structure, were relatively stable and were transported to the cytoplasm, where they could be colocalized with their target RNA. The activity of the exported ribozymes was significantly higher than that of ribozymes that remained in the nucleus. This chapter examines the methods for construction of the appropriate vector that can produce tRNA-driven ribozymes with high-level intracellular activity and for analysis of the constructs in cells.