1. Functional genomic and culture independent approaches to search for protease
activity in Forest soils in Puerto Rico
Z.M. Corales-Tirado, R.K. Rivera-Valentin, C. Rios-Velazquez, University of Puerto
Rico- Mayagüez, Biology Department.
The recognition that most microorganisms in the environment cannot be culture by
standard methods stimulated the development of metagenomics, which is the genomic
analysis of uncultured microorganisms. Modern biotechnology has a steadily increasing
demand for novel bioactive compounds, biomolecules, and enzymes. Natural and genetic
diversity of the soil metagenome has so far been the best supplier for these novel
molecules. Enzymes production is one of the most important drivers of the industrial
processing industry because it uses modern biology in the production process, and
produces the biotechnological products needed for applications in other industrial areas.
Some of these industrial enzymes are the proteases, which have important
biotechnological applications and execute a large variety of functions. The proteases
represent one of the three largest groups of industrial enzymes and have application in
detergents, leather industry, food industry, pharmaceutical industry and bioremediation
processes. Probably the largest application of proteases is in laundry detergents, where
they help removing proteins based strains from clothing. Two approaches, the function-
driven analysis and the sequence-driven analysis, have emerged to extract biological
information from metagenomic libraries. Here, we used the function-driven analysis,
which is initiated by identification of clones that express a desired trait, followed by
characterization of the active clones by sequence and biochemical analysis. We screened
three metagenomic libraries generated from forests in Puerto Rico. Serial dilutions of the
libraries were spread on Petri plates supplemented with non-fat dry milk. A clear zone
around the tested clones indicated clones with potential for protease activity. A total 20
potential clones were found which have being analyzed through biochemical and
molecular techniques to confirm whether the activity is due to the cloned fragment. The
screening of detergent proteases for industrial production has a great importance because
detergent enzymes account for about 30% of the total worldwide enzyme production, and

2. represent one of the largest and most successful applications of modern industrial
biotechnology.