Variability of nuclear mitochondrial DNA regions in the genome of autochthonous and cosmopolitan pig breeds

Insertions of mitochondrial DNA (mtDNA) into the nuclear genome have been detected in many eukaryotes. These sequences, originated by horizontal transfer of mtDNA fragments of both coding and non-coding regions, produces nuclear DNA sequences of mitochondrial origin (NUMTs). Similarly, to other inserted elements, NUMTs have been used to reconstruct the history of animal domestication and breed development at the genome level. For the same objective, in this study, we explored the potential informativity of NUMT regions’ variability in six autochthonous Italian pig breeds (Apulo Calabrese, Casertana, Cinta Senese, Mora Romagnola, Nero Siciliano and Sarda) and three Italian heavy pig breeds (Italian Landrace, Italian Large White and Italian Duroc). We initially detected NUMTs by mining the Sus scrofa reference genome (Sscrofa11.1) and several additional drafted pig genomes, including those assembled for several breeds. BLAST tool was used to map mtDNA sequences over the nuclear DNA genome. We then integrated this information by mining whole genome sequencing (WGS) data from DNA pools including several pigs or from single pigs. On the whole, about 700 NUMTs were retrieved by mining the assembled reference genome versions, 74% of which were also detected by mining WGS datasets. A total of 36 novel insertion events (de novo NUMTs, which were not reported in any assembled reference pig genome) were identified by evaluating discordant mapped reads and/or soft clipped reads. A total of 21 primer pairs, targeting six NUMTs detected from the reference genome and 15 de novo NUMTs, were designed to validate these regions by PCR and Sanger sequencing. PCR amplification was obtained in a panel of 30-35 pigs of several breeds, for a total of >300 analysed animals. As expected from the in silico analyses, we confirmed the presence at high frequency of several targeted NUMTs in all analysed breeds. A few NUMT insertions were highly frequent (or almost private) in some breeds and can be very informative in the reconstruction of the domestication process of Sus scrofa and in the definition of the genetic history of many pig breeds. Overall, this study provided an updated porcine genome map of NUMTs. We also demonstrated that several NUMTs are polymorphic in some breeds (i.e. presence/absence of the insertion) and that, for this reason, can constitute interesting markers to trace back the origin of pig populations.