Detecting clonemate pairs in multicellular diploid clonal species based on a shared heterozygosity index

Clonal reproduction, the formation of nearly identical individuals via mitosis in the absence of genetic recombination, is a very common reproductive mode across plants, fungi and animals. To detect clonal genetic structure, genetic similarity indices based on shared alleles are widely used, such as the Jaccard index, or identity by state. Here we propose a new pairwise genetic similarity index, the SH index, based on segregating genetic marker loci (typically single nucleotide polymorphisms) that are identically heterozygous for pairs of samples (N_SH ). To test our method, we analyse two old seagrass clones (Posidonia australis, estimated to be around 8500 years old; Zostera marina, >750 years old) along with two young Z. marina clones of known age (17 years old). We show that focusing on shared heterozygosity amplifies the power to distinguish sample pairs belonging to different clones compared to methods focusing on all shared alleles. Our proposed workflow can successfully detect clonemates at a location dominated by a single clone. When the collected samples involve two or more clones, the SH index shows a clear gap between clonemate pairs and interclone sample pairs. Ideally N_SH should be on the order of approximately ≥3000, a number easily achievable via restriction-site associated DNA (RAD) sequencing or whole-genome resequencing. Another potential application of the SH index is to detect possible parent-descendant pairs under selfing. Our proposed workflow takes advantage of the availability of the larger number of genetic markers in the genomic era, and improves the ability to distinguish clonemates from nonclonemates in multicellular diploid clonal species.