Fourth-degree kinship analysis using 5198 single nucleotide polymorphisms detected by nanopore sequencing platform

Nanopore sequencing technology has high portability and affordability of devices, long sequencing reads, and GB-level data yields. In forensic applications, nanopore sequencing has been used in the identification of forensic short tandem repeats (STRs) and single nucleotide polymorphisms (SNPs). Comparable to limited STRs, SNP markers have low per-locus discriminating power but are highly sufficient. Although numerous SNP panels consisting of thousands of loci have been developed, till now only ∼100 SNP panels have been evaluated using nanopore sequencing. In this study, we collected 24 DNA samples and employed a 9102 SNPs panel with hybridization capture for enrichment, followed by sequencing using Oxford Nanopore Technologies MinION device and SNP genotyping with NASTRA software. Parallel sequencing and genotyping were performed using Illumina NovaSeq, and we found the consistency of SNP genotypes with nanopore sequencing was over 99.3 %. Next, we filtered X or Y chromosomal markers, 3-6 multiple-allelic SNPs, and further excluded these failing Hardy-Weinberg Equilibrium (HWE) and Linkage Disequilibrium (LD), and obtained 5198 (57.1 %) SNPs. Based on the 5198 SNPs and likelihood ratio methods, 23 pairs of third-degree relatives were discriminated from the unrelated with a 100 % sensitivity and a 98.81 % specificity; 18 pairs of fourth-degree relatives had a 77.78 % sensitivity and 98.47 % specificity. The more close relatives were all correct in the kinship testing.