Developmental validation of a high-resolution panel genotyping 639 Y-chromosome SNP and InDel markers and its evolutionary features in Chinese populations

Abstract

Uniparental-inherited haploid genetic marker of Y-chromosome single nucleotide polymorphisms (Y-SNP) have the power to provide a deep understanding of the human evolutionary past, forensic pedigree, and bio-geographical ancestry information. Several international cross-continental or regional Y-panels instead of Y-whole sequencing have recently been developed to promote Y-tools in forensic practice. However, panels based on next-generation sequencing (NGS) explicitly developed for Chinese populations are insufficient to represent the Chinese Y-chromosome genetic diversity and complex population structures, especially for Chinese-predominant haplogroup O. We developed and validated a 639-plex panel including 633 Y-SNPs and 6 Y-Insertion/deletions, which covered 573 Y haplogroups on the Y-DNA haplogroup tree. In this panel, subgroups from haplogroup O accounted for 64.4% of total inferable haplogroups. We reported the sequencing metrics of 354 libraries sequenced with this panel, with the average sequencing depth among 226 individuals being 3,741×. We illuminated the high level of concordance, accuracy, reproducibility, and specificity of the 639-plex panel and found that 610 loci were genotyped with as little as 0.03 ng of genomic DNA in the sensitivity test. 94.05% of the 639 loci were detectable in male-female mixed DNA samples with a mix ratio of 1:500. Nearly all of the loci were genotyped correctly when no more than 25 ng/μL tannic acid, 20 ng/μL humic acid, or 37.5 μM hematin was added to the amplification mixture. More than 80% of genotypes were obtained from degraded DNA samples with a degradation index of 11.76. Individuals from the same pedigree shared identical genotypes in 11 male pedigrees. Finally, we presented the complex evolutionary history of 183 northern Chinese Hans and six other Chinese populations, and found multiple founding lineages that contributed to the northern Han Chinese gene pool. The 639-plex panel proved an efficient tool for Chinese paternal studies and forensic applications.

Keywords: Forensic genetics; Haplogroup inference; Next-generation sequencing; Single nucleotide polymorphism; Y-chromosome.

Fig. 1

Haplogroup distribution of the 639-plex panel. (A) The number of markers assigned to different haplogroups in the 639-plex panel. (B) The number of inferable haplogroups in the 639-plex panel

Fig. 2

Evaluation of the 639-plex panel by sensitivity, simulated degradation, and specificity studies. (A) Sensitivity data using series dilutions of 2800M genomic DNA. (B) Sensitivity data in male-female mixed samples. (C) Y-chromosome specificity data using commercial female genomic DNA samples. (D) Simulated degradation data of the 639-plex panel

Fig. 3

Quality scores for data in sensitivity studies. (A) Mean quality scores for the sequencing reads in sensitivity experiments. (B) FastQC profile for the sequencing reads with 0.03 ng of 2800M input DNA.

Fig. 4

PCR inhibition studies of the 639-plex panel

Fig. 5

Fine-scale genetic history of Chinese populations inferred from our developed Y-chromosome panel. (A) Network relationship of major Y-chromosome lineages observed in Liaoning Han populations. (B) Principal component analysis results among seven Chinese populations. (C) The heatmap showed the allele frequency spectrum of Chinese people. (D ~ E) Phylogenetic relationship reconstructed based on the sequence variations

Fig. 6

Haplogroup distribution and all observed derived markers associated with haplogroup O among the 183 unrelated Liaoning Han individuals. Five gradients of orange blocks represented the frequency of detected terminal haplogroups. Haplogroups above dotted lines were not covered in the panel