Directed evolution of novel polymerases.

Abstract

DNA and RNA polymerases evolved to function in specific environments with specific substrates to propagate genetic information in all living organisms. The commercial availability of these polymerases has revolutionized the biotechnology industry, but for many applications native polymerases are limited by their stability or substrate recognition. Thus, there is great interest in the directed evolution of DNA and RNA polymerases to generate enzymes with novel, desired properties, such as thermal stability, resistance to inhibitors, and altered substrate specificity. Several screening and selection approaches have been developed, both in vivo and in vitro, and have been used to evolve polymerases with a variety of important activities. Both the techniques and the evolved polymerases are reviewed here, along with a comparison of the in vivo and in vitro approaches.