The Construction Of Y-SNP Multiplex Amplification Assay And Its Application In Ancestry Inference And Paternal Pedigree Identification

Objective:The aim of this study is to screen the Y-SNP loci that have a high frequency of distribution in the East Asian population,establish a Y-SNP multiplex amplification assay and apply in ancestry inference and familal searching in forensic DNA.The genetic polymorphism of Y chromosome D-M174 haplogroup and sub-haplogroups in East Asia for the Y chromosome forensic investigations is explored.Comprehensive analysis of Y-STRs and Y-SNPs and network analysis is used in familal searching.Methods:Based on the previous research,Y-SNP loci was screened from literature and Y-DNA Haplogroup Tree on ISOGG web.Y-SNP multiplex amplification assay were constructed by mini-sequencing and used to genotype 1426 unrelated male individuals.The genotypes on Y-STR loci were obtained by DNATyperTM Y26 kit.The haplogroup frequencies and population relationships were calculated using direct counting method,heatmap,phylogenetic cluster and network.Furthermore,samples of 180 males from two pedigrees with different surnames were collected.Haplotypes at Y-STR loci were genotyped with YfilerTM Plus Kit.The genotypes of 42 Y-SNP in 36 males selected from 180 males were obtained by mini-sequencing.The haplotypes were analyzed using direct counting method and network to discuss the variations of Y-STR haplotypes within pedigree and the distribution of Y-SNP haplogroups.Results:In this study,24 Y-SNP core haplogroup and 37 Y-SNP O haplogroup multiplex amplification system were successfully established with good specificity and balance,high repeatability and accuracy.The result of haplogroup frequencies and population relationships showed that the frequencies of Y chromosome SNP haplogroup vary greatly among the populations studied.Our results revealed that D-M174 haplogroup distributed with high frequency in Tibetans(40.96%)and Japanese(35.71%),and low frequency among the surrounding area of Tibet and other East Asian populations,even no distribution.Furthermore,the distributions of Y-SNP haplogroup in the population were also different.In addition,in paternal lineage surveys,18 Y-STR haplotypes were found in 63 individuals of the P1 pedigree,and the 17 individuals selected all be-longed to the N-M231 haplogroup;35 haplotypes were found in 117 samples of the P2 pedigree,and the 19 individuals selected all belonged to Olalalal-F78 haplogroup.The network results showed that the P1 pedigree had a central tendency,while the P2 pedigree had two centers,step-by-step mutation among Y-STR haplotypes within pedigree visually and clearly showed in the network.Conclusion:The Y-SNP multiplex amplification assay can be used to genotype alleles.The distribution of Y chromosome D haplogroup in East Asian populations have obvious ethnic and geographical characteristic,and aim to provide for forensic casework.The Y-SNP information and the network method integrated with the Y-STR during pedigree investigation can largely broaden the scale in pedigree investigation,get more reliable paternal pedigree structure and increase the accuracy of paternal lineage identification.