Single-Cell/Fragment DNA-Seq of Rhesus Embryos

A major contributor to in vitro fertilization (IVF) failure and embryo loss is aneuploidy. Aneuploid human pre-implantation embryos often sequester missegregated chromosomes into micronuclei during mitotic divisions, while concurrently undergoing a process called cellular fragmentation. Given the coordinated timing of these two events, we hypothesized that cellular fragmentation is a response to the encapsulation of missegregated chromosomes within micronuclei. To test this hypothesis, embryos from the closely-related rhesus monkey were comprehensively examined using single-cell/-fragment DNA-sequencing (scDNA-Seq) for copy number variation (CNV) and single nucleotide polymorphism (SNP) analyses. Cleavage-stage embryos were disassembled into single blastomeres, cellular fragments, and polar bodies and individually collected for low coverage sequencing. Additionally, genomic DNA from the rhesus parental donors of sperm and oocytes used for embryo production were also sequenced for SNP parentage analysis. Male and female euploid rhesus skin fibroblasts (AG08312 and AG08305, Coriell Institute, Camden, NJ) ranging in number from 1 cell to 10 cells were used for CNV pipeline development. The accuracy of the CNV pipeline for calling trisomies and monosomies was tested with single human fibroblasts carrying known aneuploidies (GM10179 and AG05024, Coriell Institute, Camden, NJ). Our findings show that whole and partial chromosomes originating from either parent can be eliminated from the embryo through cellular fragmentation. We also demonstrate that copy number and ploidy states amongst blastomeres in cleavage-stage embryos range in complexity, but in some cases, embryos can overcome these chromosomal errors through exclusion of chaotic blastomeres.