Mitochondrial nucleic acid containing mitochondrial DNA (mtDNA) was prepared from the purified mitochondria by protease K treatment, phenol extraction, chloroform/isoamyl-alcohol extraction and ethanol precipitation, followed by linearization of mtDNA by restriction enzyme digestion. The mtDNA and spike-in lambda DNA were treated with 9.2M bisulfite, and used for PBAT library preparation as a template.