Genomic libraries derived from environmental DNA (metagenomic libraries) are useful for characterizing
uncultured microorganisms. However, conventional library-screening techniques permit characterization of
relatively few environmental clones. Here we describe a novel approach for characterization of a metagenomic
library by hybridizing the library with DNA from a set of groundwater isolates, reference strains, and communities.
A cosmid library derived from a microcosm of groundwater microorganisms was used to construct
a microarray (COSMO) containing ~1-kb PCR products amplified from the inserts of 672 cosmids plus a set
of 16S ribosomal DNA controls. COSMO was hybridized with Cy5-labeled genomic DNA from each bacterial
strain, and the results were compared with the results for a common Cy3-labeled reference DNA sample
consisting of a composite of genomic DNA from multiple species. The accuracy of the results was confirmed by
the preferential hybridization of each strain to its corresponding rDNA probe. Cosmid clones were identified
that hybridized specifically to each of 10 microcosm isolates, and other clones produced positive results with
multiple related species, which is indicative of conserved genes. Many clones did not hybridize to any
microcosm isolate; however, some of these clones hybridized to community genomic DNA, suggesting that they
were derived from microbes that we failed to isolate in pure culture. Based on identification of genes by end
sequencing of 17 such clones, DNA could be assigned to functions that have potential ecological importance,
including hydrogen oxidation, nitrate reduction, and transposition. Metagenomic profiling offers an effective
approach for rapidly characterizing many clones and identifying the clones corresponding to unidentified
species of microorganisms
