# A non-hypothesis-driven practical laboratory activity on functional metagenomics: “fishing” protein-coding DNA sequences from microbiomes

## TL;DR

This paper introduces a hands-on lab activity for functional metagenomics, allowing students to construct and analyze DNA libraries from microbiomes to explore gene functions.

## Contribution

A practical, student-tested protocol for building metagenomic DNA libraries from intron-less genomes for educational and research purposes.

## Key findings

- The protocol enables rapid library construction from bacterial or phage DNA for functional analysis.
- The approach supports training in bioinformatics and proteomics techniques for gene validation.
- The method offers flexibility for diverse research projects by varying biological sources and validation methods.

## Abstract

Practical laboratory of the most functional metagenomics courses focuses on activities aimed at providing specific skills in bioinformatics through the analysis of genomic datasets. However, sequence-based analyses of metagenomes should be complemented by function-based analyses, to provide evidential knowledge of gene function. A “true” functional metagenomic approach relies on the construction and screening of metagenomic libraries - physical libraries that contain DNA cloned from metagenomes of various origin. The information obtained from functional metagenomics will help in future annotations of gene function and serve as a complement to sequence-based metagenomics. Here, we describe a simple protocol for the construction of a metagenomic DNA library, optimized and tested by a team of undergraduate biotechnology students. This protocol is based on a technique developed in our laboratory and currently used for research. Using this protocol, libraries of protein domains can be quickly generated, from the DNA of any intron-less genome, such as those of bacteria or phages. Therefore, these libraries provide a valuable platform for training students in various validation tools, including computational methods - for example, metagenome assembly, functional annotation - and proteomics techniques, including protein expression and analysis. By varying the biological source and validation pipeline, this approach offers virtually limitless opportunities for innovative thesis research projects.