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Directed Enzyme Evolution - Screening and Selection Methods (Paperback, Softcover reprint of hardcover 1st ed. 2003)
Loot Price: R3,337
Discovery Miles 33 370
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Directed Enzyme Evolution - Screening and Selection Methods (Paperback, Softcover reprint of hardcover 1st ed. 2003)
Series: Methods in Molecular Biology, 230
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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. In many ways, the second step is the most
challenging. No matter how cleverly designed or diverse the
starting library, without an effective screening strategy the
ability to isolate useful clones is severely diminished. The best
screens are (1) high throughput, to increase the likelihood that
useful clones will be found; (2) sufficiently sen- tive (i. e. ,
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. 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. " The protocols in Directed Enzyme Evolution
describe a series of detailed p- cedures of proven utility for
directed evolution purposes. The volume begins with several
selection strategies for enzyme evolution and continues with assay
methods that can be used to screen enzyme libraries. Genetic
selections offer the advantage that functional proteins can be
isolated from very large libraries s- ply by growing a population
of cells under selective conditions.
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