|Test Type||Callable Loci||Poznik Loci||Poor Alignment||Total Loci||Samples (n)||Est. Years/SNP||Histogram|
The coverage table is produced using BWA-MEM aligned BAMs on the GRCh38 reference genome. The BAMs are processed by GATK's CallableLoci tool with default settings. For a location to be considered callable it must have four reads overlapping the site. No more than ten percent of those reads may have a PHRED-scaled alignment quality of less than 10. Yielding a heuristic combined quality indicating less than 0.01% chance all the reads are aligned incorrectly to the site.
Callable - The total number of sites sequenced with 4 or more reads. Each read has less than 10% likelihood of being misaligned.
Poznik Loci - The callable sites occurring Poznik et al. (2013), Sequencing Y Chromosomes Resolves Discrepancy in Time to Common Ancestor of Males Versus Females. Retrieved Feb 17, 2018 from Science. These regions are a superset of the combBED used by Adamov's group. The sites include areas outside of Big Y's targetted sequencing regions.
Poor Alignment - The sites having more than four reads, but the aligner indicates a high likelihood the results may not be correct.
This tool has revealed a large coefficient of variance in 10x Genomics Chromium genome sequencing. These variances are due to poor sample quality and highlight the importance of following collection protocols and quality screening. While several examples support the case saliva samples on this technology are viable, just as many are showing disappointing results. It would seem blood samples are needed to fully realize the potential of this test and mitigate risks unless additional quality gates are put in check prior to sequencing.
The vendor notes that they invested in additional sequencing for the customers impacted by this issue at no charge. The problem is at least one sample was so badly contaminated a completely new run would be required. When viewed with two reads as miminum for calling instead of four, this sample could be compared with the other men in his cluster but at a lower specificity.