![]() ![]() ” The protocols in Directed Enzyme Evolution describe a series of detailed p- cedures of proven utility for directed evolution purposes. Regarding this last point, almost anyone who has attempted a directed evolution experiment has learned firsthand the truth of the dictum “you get what you screen for. , good signal to noise) to allow the isolation of lower activity clones early in evolution (3) sufficiently reproducible to allow one to find small improvements (4) robust, which means that the signal afforded by active clones is not dependent on difficult-to-control environmental variables and, most importantly, (5) sensitive to the desired function. The best screens are (1) high throughput, to increase the likelihood that useful clones will be found (2) sufficiently sen- tive (i. ![]() No matter how cleverly designed or diverse the starting library, without an effective screening strategy the ability to isolate useful clones is severely diminished. In many ways, the second step is the most challenging. Directed evolution comprises two distinct steps that are typically applied in an iterative fashion: (1) generating molecular diversity and (2) finding among the ensemble of mutant sequences those proteins that perform the desired fu- tion according to the specified criteria.
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