Towards the detection of copy number variation from single sperm sequencing in cattle.

BACKGROUND: Copy number variation (CNV) has been routinely studied using bulk-cell sequencing. However, CNV is not well studied on the single-cell level except for humans and a few model organisms. RESULTS: We sequenced 143 single sperms of two Holstein bulls, from which we predicted CNV events using 14 single sperms with deep sequencing. We then compared the CNV results derived from single sperms with the bulk-cell sequencing of one bull's family trio of diploid genomes. As a known CNV hotspot, segmental duplications were also predicted using the bovine ARS-UCD1.2 genome. Although the trio CNVs validated only some single sperm CNVs, they still showed a distal chromosomal distribution pattern and significant associations with segmental duplications and satellite repeats. CONCLUSION: Our preliminary results pointed out future research directions and highlighted the importance of uniform whole genome amplification, deep sequence coverage, and dedicated software pipelines for CNV detection using single cell sequencing data.

Genome-wide recombination map construction from single sperm sequencing in cattle.

BACKGROUND: Meiotic recombination is one of the important phenomena contributing to gamete genome diversity. However, except for human and a few model organisms, it is not well studied in livestock, including cattle. RESULTS: To investigate their distributions in the cattle sperm genome, we sequenced 143 single sperms from two Holstein bulls. We mapped meiotic recombination events at high resolution based on phased heterozygous single nucleotide polymorphism (SNP). In the absence of evolutionary selection pressure in fertilization and survival, recombination events in sperm are enriched near distal chromosomal ends, revealing that such a pattern is intrinsic to the molecular mechanism of meiosis. Furthermore, we further validated these findings in single sperms with results derived from sequencing its family trio of diploid genomes and our previous studies of recombination in cattle. CONCLUSIONS: To our knowledge, this is the first large-scale single sperm whole-genome sequencing effort in livestock, which provided useful information for future studies of recombination, genome instability, and male infertility.

Construction of PRDM9 allele-specific recombination maps in cattle using large-scale pedigree analysis and genome-wide single sperm genomics.

PRDM9 contributes to hybrid sterility and species evolution. However, its role is to be confirmed in cattle, a major domesticated livestock species. We previously found an association near PRDM9 with cattle recombination features, but the causative variants are still unknown. Using millions of genotyped cattle with pedigree information, we characterized five PRDM9 alleles and generated allele-specific recombination maps. By examining allele-specific recombination patterns, we observed the impact of PRDM9 on global distribution of recombination, especially in the two ends of chromosomes. We also showed strong associations between recombination hotspot regions and functional mutations within PRDM9 zinc finger domain. More importantly, we found one allele of PRDM9 to be very different from others in both protein composition and recombination landscape, indicating the causative role of this allele on the association between PRDM9 and cattle recombination. When comparing recombination maps from sperm and pedigree data, we observed similar genome-wide recombination patterns, validating the quality of pedigree-based results. Collectively, these evidence supported PRDM9 alleles as causal variants for the reported association with cattle recombination. Our study comprehensively surveyed the bovine PRDM9 alleles, generated allele-specific recombination maps, and expanded our understanding of the role of PRDM9 on genome distribution of recombination.