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 - 1x minimum average genomic coverage per analysed pair of individuals between. Here we present an updated version of a method that enables estimates of 1st and 2nd-degrees of relatedness with as little as 0.026x average coverage, or around 1.3 million aligned reads per sample - 4 times less data than 0.1x. By using simulated data to estimate false positive error rates, we further show that a threshold even as low as 0.012x, or around 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 previously excluded from 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.