Investigating human mitochondrial genomes in single cells

Mitochondria host multiple copies of their own small circular genome that has been extensively studied to trace the evolution of the modern eukaryotic cell and discover important mutations linked to inherited diseases. Whole genome and exome sequencing have enabled the study of mtDNA in a large number of samples and experimental conditions at single nucleotide resolution, allowing the deciphering of the relationship between inherited mutations and phenotypes and the identification of acquired mtDNA mutations in classical mitochondrial diseases as well as in chronic disorders, ageing and cancer. By applying an ad hoc computational pipeline based on our MToolBox software, we reconstructed mtDNA genomes in single cells using whole genome and exome sequencing data obtained by different amplification methodologies (eWGA, DOP-PCR, MALBAC, MDA) as well as data from single cell Assay for Transposase Accessible Chromatin with high-throughput sequencing (scATAC-seq) in which mtDNA sequences are expected as a byproduct of the technology. We show that assembled mtDNAs, with the exception of those reconstructed by MALBAC and DOP-PCR methods, are quite uniform and suitable for genomic investigations, enabling the study of various biological processes related to cellular heterogeneity such as tumor evolution, neural somatic mosaicism and embryonic development.