PT - JOURNAL ARTICLE AU - Anitha D Jayaprakash AU - Erica Benson AU - Raymond Liang AU - Jaehee Shim AU - Luca Lambertini AU - Mike Wigler AU - Stuart A Aaronson AU - Ravi Sachidanandam TI - Mito-seek enables deep analysis of mitochondrial DNA, revealing ubiquitous, stable heteroplasmy maintained by intercellular exchange AID - 10.1101/007005 DP - 2014 Jan 01 TA - bioRxiv PG - 007005 4099 - http://biorxiv.org/content/early/2014/07/10/007005.short 4100 - http://biorxiv.org/content/early/2014/07/10/007005.full AB - Eukaryotic cells carry two genomes, nuclear (nDNA) and mitochondrial (mtDNA), which are ostensibly decoupled in their replication, segregation and inheritance. It is increasingly appreciated that heteroplasmy, the occurrence of multiple mtDNA haplotypes in a cell, plays an important biological role, but its features are not well understood. Until now, accurately determining the diversity of mtDNA has been difficult due to the relatively small amount of mtDNA in each cell (< 1% of the total DNA), the intercellular variability of mtDNA content and copies of mtDNA pseudogenes in nDNA. To understand the nature of heteroplasmy, we developed Mito-seek, a novel technique that purifies and sequences mtDNA. Mito-seek yields high purity (> 98%) mtDNA and its ability to detect rare variants is limited only by sequencing depth, providing unprecedented sensitivity and specificity. Using Mito-seek, we confirmed the ubiquity of heteroplasmy by analyzing mtDNA from a diverse set of cell lines and human samples. By applying Mito-seek to colonies derived from single cells, we showed that heteroplasmy is stably maintained in individual daughter cells over multiple cell divisions. Our simulations indicate that the stability of heteroplasmy can be facilitated by the exchange of mtDNA between cells. We also explicitly demonstrate this exchange by co-culturing cell lines with distinct mtDNA haplotypes. Our results shed new light on the maintenance of heteroplasmy and provide a novel platform to investigate various features of heteroplasmy in normal and diseased tissues.