TKGWV2: an ancient DNA relatedness pipeline for ultra-low coverage whole genome shotgun data

Estimation of genetically related individuals is playing an increasingly important role in the ancient DNA field. In recent years, the numbers of sequenced individuals from single sites have been increasing, reflecting a growing interest in understanding the familial and social organisation of ancient populations. Although a few different methods have been specifically developed for ancient DNA, namely to tackle issues such as low-coverage homozygous data, they require a 0.1–1× minimum average genomic coverage per analysed pair of individuals. Here we present an updated version of a method that enables estimates of 1st and 2nd-degrees of relatedness with as little as 0.026× average coverage, or around 18,000 SNPs from 1.3 million aligned reads per sample with average length of 62 bp—four times less data than 0.1× coverage at similar read lengths. By using simulated data to estimate false positive error rates, we further show that a threshold even as low as 0.012×, or around 4000 SNPs from 600,000 reads, will always show 1st-degree relationships as related. Lastly, by applying this method to published data, we are able to identify previously undocumented relationships using individuals that had been excluded from prior kinship analysis due to their very low coverage. This methodological improvement has the potential to enable relatedness estimation on ancient whole genome shotgun data during routine low-coverage screening, and therefore improve project management when decisions need to be made on which individuals are to be further sequenced.