Numerous insertions of mitochondrial DNA in the genome of the northern mole vole, Ellobius talpinus

Abstract

Background Ellobius talpinus is a subterranean rodent representing an attractive model in population ecology studies due to its highly special lifestyle and sociality. In such studies, mitochondrial DNA (mtDNA) is widely used. However, if nuclear copies of mtDNA, aka NUMTs, are present, they may co-amplify with the target mtDNA fragment, generating misleading results. The aim of this study was to determine whether NUMTs are present in E. talpinus. Methods and results PCR amplification of the putative mtDNA CytB-D-loop fragment using ‘universal’ primers from 56 E. talpinus samples produced multiple double peaks in 90% of the sequencing chromatograms. To reveal NUMTs, molecular cloning and sequencing of PCR products of three specimens was conducted, followed by phylogenetic analysis. The pseudogene nature of three out of the seven detected haplotypes was confirmed by their basal positions in relation to other Ellobius haplotypes in the phylogenetic tree. Additionally, ‘haplotype B’ was basal in relation to other E. talpinus haplotypes and found present in very distant sampling sites. BLASTN search revealed 195 NUMTs in the E. talpinus nuclear genome, including fragments of all four PCR amplified pseudogenes. Although the majority of the NUMTs studied were short, the entire mtDNA had copies in the nuclear genome. The most numerous NUMTs were found for rrnL, COXI, and D-loop. Conclusions Numerous NUMTs are present in E. talpinus and can be difficult to discriminate against mtDNA sequences. Thus, in future population or phylogenetic studies in E. talpinus, the possibility of cryptic NUMTs amplification should always be taken into account.

Repeat-rich regions cause false positive detection of NUMTs: a case study in amphibians using an improved cane toad reference genome

“…For example, NUMTs can lead to an overestimation of the number of novel mutations on a population scale, potentially resulting in inaccurate inferences related to allelic frequency, population structure, demography, and phylogenetic relationships, or misidentification of heteroplasmy (Maude, et al 2019;Schultz and Hebert 2022;Song, et al 2008). Methods to detect NUMTs include bioinformatic prediction (Dayama, et al 2014;Dayama, et al 2020;Hebert, et al 2023;) and laboratory-based detection (PCR amplifications) (Kuprina, et al 2023;Machida and Lin 2017), but their implementation is often restricted by cost and efficiency.…”