Mitochondrial Dna Heteroplasmy Type In Inbred Mice

Inbred mouse strains are widely used to identify gene candidates to account for phenotypic variability for various traits, including endurance trainability. While the major mitochondrial DNA (mtDNA) sequence is known for most strains, changes in sequence across mtDNA copies, or heteroplasmy, have yet to be well-defined or completely understood. Therefore, a more thorough understanding of heteroplasmy would augment our understanding of the role of mitochondrial genetics in mediating the individual adaptive response to endurance training. PURPOSE: This preliminary investigation will characterize types of mtDNA variants and heteroplasmy as determined by comparison of the reference strain (C57BL/6 J mouse) to a battery of mouse strains. METHODS: DNA was isolated from the lungs of 32 inbred male mice (n = 2 mice/strain), and mtDNA was amplified by long range PCR, then tagged with Nextera libraries and sequenced on a MiSeq instrument. A site was considered heteroplasmic when the minor allele frequency was ≥1% across the mtDNA copies. We determined the mean heteroplasmy frequency (number of heteroplasmic sites) for each strain, then characterized each heteroplasmic site as either Type 1 or Type 2 heteroplasmy based on the presence of heteroplasmy in either the alternate (Type 1) or reference allele (Type 2). RESULTS: Overall, we found Type 1 heteroplasmies occurred in 30 of the 32 strains, while 7 of the 32 strains had Type 2 heteroplasmy. NZO/HILtJ (198 sites), C3H/HeOuJ (29 sites), and DBA/2 J (19 sites) strains showed the highest frequency Type 1 heteroplasmy, while MOLF/EiJ (356 sites), NZO/HILtJ (81 sites), and PWK/J (71 sites) strains demonstrated the highest frequency Type 2 heteroplasmy. Additionally, analysis of respiratory chain encoding genes revealed a substantial frequency of Type 2 heteroplasmy in the mt-ND5 gene of MOLF/EiJ (6 sites), PWD/PhJ (6 sites), and PWK/J (7 sites) strains, while the NZO strain showed no presence of Type 2 heteroplasmy in the mt-ND5 gene. CONCLUSION: This is the first study in which heteroplasmy has been characterized as occurring in either the reference or alternate allele. Future work in our lab will explore potential relationships between endurance trainability and heteroplasmy type. ACKNOWLEDGEMENTS: