Dissection of uniparental reproduction barriers in mice using haploid embryonic stem cells [RRBS]

Uniparental reproduction is widespread among lower vertebrates, but not in mammals. Researchers have produced bimaternal mice with a deletion of the H19 imprinted region. However, the mechanism of a single deletion in an immature oocyte sufficient to cross uniparental barriers is unknown. We found bimaternal-derived mice to be defective in multiple aspects. More importantly, bipaternal reproduction has not been achieved in mammals. Thus, the barrier of uniparental reproduction in mammals has not been elucidated. We identified a DNA hypomethylation status in haploid embryonic stem cells similar to that in primordial germ cells, with which bimaternal and bipaternal mice can be obtained by injection of haploid ES cells with genetic modifications. The phenotypic analyses of derived mice support the genetic conflict theory of genomic imprinting. Taken together, our results highlight the factors necessary for crossing uniparental reproduction barriers in mammals. Reduced representation bisulfite sequencing (RRBS) for a global DNA methylation analysis of wild type (WT) or the differentially methylated region knocking out (DMR-KO) haploid ESCs androgenetic haploid ESCs (ahESC), parthenogenetic haploid ESCs (phESCs), diploid ESCs, embryos and brain tissue. Compared to wild type (WT) embryo, which was produced by normal zygote deriving from normal sperm and oocyte, the bimaternal (BM) embryo was produced through injecting a phESC into an MII oocyte and the bipaternal (BP) embryo was produced through injecting injecting an ahESC and a sperm into an enucleated oocyte.