Directed evolution, the laboratory process by which biological entities with desired traits are created through iterative rounds of genetic diversification and library screening or selection, has become one of the most useful and widespread tools in basic and applied biology. From its roots in classical strain engineering and adaptive evolution, modern directed evolution came of age 20 years ago with the demonstration of repeated rounds of polymerase chain reaction (PCR)-driven random mutagenesis and activity screening to improve protein properties. Since then, numerous techniques have been developed that have enabled the evolution of virtually any protein, pathway, network, or entire organism of interest. Here, we recount some of the major milestones in the history of directed evolution, highlight the most promising recent developments in the field, and discuss the future challenges and opportunities that lie ahead. © 2013 American Institute of Chemical Engineers AIChE J, 59: 1432–1440, 2013
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