Patent ID: 8557742
Filing Date: 2013-10-15
Classification: C12N

Abstract:
1. A selection method for enhanced enzyme activity on an insoluble substrate, the method comprising: (a) providing a plurality of polynucleotides encoding variants of one or more enzyme(s) that act(s) on an insoluble substrate, wherein the plurality of polynucleotides is linked to a plurality of solid phases and the linker or the solid phases comprise a substrate for the one or more enzyme(s); (b) suspending the polynucleotide-linked solid phases in an aqueous phase comprising components for in vitro transcription/translation; (c) forming a water-in-oil emulsion, wherein the polynucleotide-linked solid phases are compartmentalized in aqueous droplets in an oil continuous phase; (d) carrying out in vitro transcription/translation to express enzyme variants within aqueous droplets of the emulsion, wherein an active enzyme variant in an aqueous droplet releases the polynucleotide(s) from the solid phase-in that droplet; and (e) separating the aqueous phase from the solid and oil phases to recover polynucleotides in the aqueous phase that have been released from the solid phases.