| The purpose of the present study was to develop new methods for the construction and screening of soil metagenomic libraries for the discovery of new antifungal and anticancer agents. Methods for DNA isolation were optimized using local soil, sediment, and sludge samples collected from Indian Creek, Beaver Dam Creek Swamp, and an anaerobic digestor, respectively. For metagenomic library construction and screening, environmental DNA was isolated from epiphytic soil collected from the Monteverde Cloud Forest Preserve in Costa Rica. The metagenomic DNA was cloned into a fosmid vector to construct a 10,000-clone library in Escherichia coli, yielding recombinant plasmids with an average size of approximately 34 kb. Three transfectants exhibited greater than 20% inhibition of respiration of MCF-7 human breast adenocarcinoma cells, and one exhibited greater than 20% growth inhibition of Candida albicans. It is clear that the methods described represent a viable approach for the identification of potentially novel bioactive compounds. |
