Growth selection method

Abstract

The invention provides growth selection methods for screening an environmental or a mixed population library of nucleic acids comprising uncharacterized nucleic acids for the presence of a nucleic acid encoding an enzyme of interest. In one aspect, methods for screening and identification of enzymes, e.g., transaminases, nitrilases, aldolases, epoxide hydrolases are provided. Methods for the production and screening of gene libraries generated from nucleic acids isolated from more than one organism for enzymes of interest, e.g., transaminases, nitrilases, aldolases, epoxide hydrolases, are also provided.

1 . A method for screening an environmental or a mixed population library of nucleic acids comprising uncharacterized nucleic acids for the presence of a nucleic acid encoding an enzyme of interest, comprising:
 (a) providing an environmental or mixed population nucleic acid library comprising a plurality of uncharacterized polypeptide-encoding nucleic acids;   (b) providing a plurality of cells, wherein the cells lack an active form of the enzyme and lack a factor essential for growth of the cells, wherein the factor is necessary to induce growth of the cells;   (c) inserting the nucleic acid library of step (a) into the cells of step (b) and incubating the cells under conditions wherein polypeptides encoded by the uncharacterized polypeptide-encoding nucleic acids of step (a) are expressed;   (d) incubating the cells in a growth medium lacking the factor necessary for cell growth; and   (e) screening the cells for growth, wherein growth of a cell identifies the presence in that cell of a nucleic acid encoding an enzyme of interest.