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Item does not contain fulltext There is an increasing availability of complete or draft genome sequences for microbial organisms. These data form a potentially valuable resource for genotype-phenotype association and gene function prediction, provided that phenotypes are consistently annotated for all the sequenced strains. In this review, we address the requirements for successful gene-trait matching. We outline a basic protocol for microbial functional genomics, including genome assembly, annotation of genotypes (including single nucleotide polymorphisms, orthologous groups and prophages), data pre-processing, genotype-phenotype association, visualization and interpretation of results. The methodologies for association described herein can be applied to other data types, opening up possibilities to analyze transcriptome-phenotype associations, and correlate microbial population structure or activity, as measured by metagenomics, to environmental parameters.