HIGH THROUGHPUT DISCOVERY OF NEW GENES FROM COMPLEX MIXTURES OF ENVIRONMENTAL MICROBES

摘要

Compositions and methods for isolating new variants of known gene sequences are provided. The methods find use in identifying variants, particularly homologs, in complex mixtures. Compositions comprise hybridization baits that hybridize to gene families of interest, particularly agricultural interest, in order to selectively enrich the polynucleotides of interest from complex mixtures. Bait sequences may be specific for a number of genes from distinct gene families of interest and may be designed to cover each gene of interest by at least 2-fold. Thus methods disclosed herein are drawn to an oligonucleotide hybridization gene capture approach for identification of new genes of interest from environmental samples. This approach bypasses the need for labor-intensive microbial strain isolation, permits simultaneous discovery of genes from multiple gene families of interest, and increases the potential to discover genes from low-abundance and unculturable organisms present in complex mixtures of environmental microbes.

主权项

1. A method for identifying genes of interest, or variants thereof, comprising:
a) obtaining complex sample, comprising said genes of interest, or variants thereof; b) preparing DNA from said complex sample for hybridization; c) mixing said prepared sample DNA with labeled baits specific for fragments of said genes of interest; d) hybridizing said labeled baits with said prepared sample DNA under conditions that allow for hybridization of said labeled baits with the corresponding gene fragments of interest to form a hybridization complex; e) sequencing said hybridized gene fragments of interest; and f) analyzing said sequences to identify said genes of interest, or variants thereof.