The phylogenetic relationship among the three genera of the family Streptomycetaceae was examined using the small and large subunit ribosomal RNA genes, and the gyrB, rpoB, trpB, atpD and recA genes. The total stretches of the analyzed ribosomal genes were 4.2kb, and those of five protein coding genes were 4.5 kb. The resultant phylogenetic trees confirmed that each genus formed an independent clade in the majority of cases. The G+C contents of rRNA genes were 56.9-58.9 mol%, and those of protein coding genes were 65.4-72.4 mol%, the latter being closer to those of the genomic DNAs. The average nucleotide sequence identity between the organisms were 94.1-96.4% for rRNA genes and 85.7-90.6% for protein coding genes, thus indicating that protein coding genes can give higher resolution than rRNA genes. In addition, the protein coding gene trees were more stable than the rRNA gene trees, supported by higher bootstrap values and other treeing algorithms. Moreover, the genome data of six Streptomyces species indicated that many protein coding genes exhibited higher correlations with genome relatedness. The combined gene sequences were also shown to give a better resolution with higher stability than any single genes, though not necessarily more correlated with genome relatedness. It is evident from this study that the rRNA gene based phylogeny can be misleading, and also that protein coding genes have a number of advantages over the rRNA genes as the phylogenetic markers including a high correlation with the genome relatedness.