Flavobacterium columnare is a Gram negative fish pathogen that causes columnaris disease, which infects populations of wild and cultured fish species. However, pathogenic mechanisms of F. columnare are largely unknown. The purpose of this research is to obtain the complete sequence of the F. columnare ATCC 49512 genome to advance pathogenesis research and increase our understanding of this pathogen. To accomplish this, genome sequencing by using Sanger and 454 sequencing was conducted. The sequences were assembled, gaps were filled, and the circular genome was autoannotated. The F. columnare genome size is 3.2 Mb and AT rich (68.5% AT). It contains 2,882 predicted proteins, 71 tRNA genes and five ribosomal RNA operons. More than half (57.1%) of the open reading frames have assigned function, which included chondroitin AC lyase, proteases, collagenases, and genes involved in biofilm formation, secretion systems, iron acquisition, and gliding motility.