Long-read-based single sperm genome sequencing for chromosome-wide haplotype phasing of both SNPs and SVs

Abstract

Abstract Although localized haploid phasing can be achieved using long read genome sequencing without parental data, reliable chromosome-scale phasing remains a great challenge. Given that sperm is a natural haploid cell, single-sperm genome sequencing can provide a chromosome-wide phase signal. Due to the limitation of read length, current short-read-based single-sperm genome sequencing methods can only achieve SNP haplotyping and come with difficulties in detecting and haplotyping structural variations (SVs) in complex genomic regions. To overcome these limitations, we developed a long-read-based single-sperm genome sequencing method and a corresponding data analysis pipeline that can accurately identify crossover events and chromosomal level aneuploidies in single sperm and efficiently detect SVs within individual sperm cells. Importantly, without parental genome information, our method can accurately conduct de novo phasing of heterozygous SVs as well as SNPs from male individuals at the whole chromosome scale. The accuracy for phasing of SVs was as high as 98.59% using 100 single sperm cells, and the accuracy for phasing of SNPs was as high as 99.95%. Additionally, our method reliably enabled deduction of the repeat expansions of haplotype-resolved STRs/VNTRs in single sperm cells. Our method provides a new opportunity for studying haplotype-related genetics in mammals.