Next generation sequencing (NGS) made it possible to provide whole genome sequences of pathogenic and commercially significant organisms in limited time, and with minimal cost. Computational comparative genomics is necessary, given that we sequence thousands of organisms every day, but our follow-up knowledge is still very limited. Nevertheless, genomic information from a single genome is insufficient to provide insights into the life style and extended view of the gene pool of a species. Multiple genomes could enrich our understanding of the relatedness of, and variations in organisms. Consequently, comparative genomic analysis remains powerful tools for identifying the orthologous genes in species, presence and absence of specific genes, evolutionary signals, and candidate
regions associated with pathogenicity. Furthermore, pangenomic strategies, together with subtractive genomics, help in highlighting the inter- and intra-species relationships, conserved core and, pan-genome for characterizing virulence factors, drug targets and vaccine candidates. In this article, we present an overview of microbial comparative genomics pre-requisites: sequencing technologies, alignment tools, annotation pipelines, databases and resources, visualization and comparative genomic tools, and strategies. Finally, we present comparative genomic and functional analysis based insights and recent findings in genus Corynebacterium.