Probing meiotic recombination and aneuploidy of single sperm cells by whole-genome sequencing

Abstract

Meiotic recombination creates genetic diversity and ensures segregation of homologous chromosomes. Previous population analyses yielded results averaged among individuals and affected by evolutionary pressures. We sequenced 99 sperm from an Asian male by using the newly developed amplification method-multiple annealing and looping-based amplification cycles-to phase the personal genome and map recombination events at high resolution, which are nonuniformly distributed across the genome in the absence of selection pressure. The paucity of recombination near transcription start sites observed in individual sperm indicates that such a phenomenon is intrinsic to the molecular mechanism of meiosis. Interestingly, a decreased crossover frequency combined with an increase of autosomal aneuploidy is observable on a global per-sperm basis.

Figure 1

Principle of whole genome phasing of an individual using the SNP linkage information from individual sperm cells. (A) We sequenced the diploid genome and identified five heterozygous SNPs with unknown linkage information shown in purple. Individual sperm cells were sequenced after MALBAC amplification, from which SNP linkage information in each sperm was used to infer the phase information in the diploid genome (B) Performance of whole genome phasing by SNP linkage in sperm cells.

Figure 2

Identifying crossover positions in individual sperm cells (A) Parental haplotype contributions are determined by comparing the percentage of reads covering the paternal or maternal SNPs, and crossover positions are detected by identifying the crossing locations of the two parental haplotypes by a hidden Markov model. (B) Resolution of crossover determination. ~60% of the crossovers can be determined within intervals of 50kb. (C) Distribution of recombination rate relative to transcription start sites (TSS).

Figure 3

Genome-wide distribution of recombination (A) Comparison of the sperm recombination rates to the HapMap and deCODE (male-specific) genetic maps across the human genome. We used a 3Mb statistical window size and a 1Mb moving step. (B) A personal genetic map. Relations of physical and genetic length of selected chromosomes. (C) Distance distribution of coexisted crossovers on the same chromosome. The experimental data is fitted with gamma distribution (α=3.35), indicating a strong deviation from random distribution. In comparison, we generated random crossovers based on physical and genetic distances.

Figure 4

Detecting aneuploidy and crossover in the same sperm. (A) Two of the four sperm cells that exhibit autosomal abnormality. Few reads are mapped to chr19 in S39, indicating a loss of chr19. Both parental haplotypes are found in chr6 of S65, indicating a disomy chr6 in the sperm. The detailed coverage analysis on all four aneuploid sperms is shown in Fig S2. (B) Distribution of the autosomal crossover number. Blue arrows indicate the number of crossover in sperm cells with autosomal aneuploidy.