The Myc oncoprotein is a transcription factor involved in a variety of human cancers. Overexpression of Myc is associated with malignant transformation. In normal cells, Myc is induced by mitotic signals, and in turn, it regulates the expression of downstream target genes. Although diverse roles of Myc have been predicted from many previous studies, detailed functions of Myc targets are still unclear. By combining chromatin immunoprecipitation (ChIP) and promoter microarrays, we identified a total of 1469 Myc direct target genes, the majority of which are novel, in HeLa cells and human primary fibroblasts. We observed dramatic changes of Myc occupancy at its target promoters in foreskin fibroblasts in response to serum stimulation. Among the targets of Myc, 107 were nuclear encoded genes involved in mitochondrial biogenesis. Genes with important roles in mitochondrial replication and biogenesis, such as POLG, POLG2, and NRF1 were identified as direct targets of Myc, confirming a direct role for Myc in regulating mitochondrial biogenesis. Analysis of target promoter sequences revealed a strong preference for Myc occupancy at promoters containing one of several described consensus sequences, CACGTG, in vivo. This study thus sheds light on the transcriptional regulatory networks mediated by Myc in vivo.