DNA methylomes in laser microdissection (LMD) captured apomictic cells to delineate nucellar embryogenesis initiation of citrus. citrus

Citrus nucellar polyembryony (NPE) is a mode of sporophytic apomixis that allows clonal propagation of rootstocks, but it impedes citrus cross breeding. To dissect the DNA methylation regulation involved in nucellar poly-embryony (NPE) in citrus, we compared the DNA methylome profiles in LMD-captured citrus apomictic cells and the nucellar sporophytic cells at two nucellar embryogenesis initiation (NEI) stages. At earlier NEI stage (3 d before anthesis), when the sexual embryo sac is undergoing mega-gametogenesis and the NEI cells are not observable under microscope, the 6-7 layers of nucellar tissues (i.e., Poly-NC and Mono-NC) were captured from the ovary sections of the poly- and mono-embryonic F1 progenies of Hirado Buntan pummelo (mono-embryonic) and Fairchild mandarin (poly-embryonic), respectively. At later NEI stage (7 d after anthesis), when the zygote in embryo sac is undergoing mitosis and the densely cytoplasmic NEI cells are observable under microscope, the NEI cells and sporophytic nucellar (SO) cells were captured from the ovary sections of the highly poly-embryonic Ponkan mandarin. In our results, The global DNA methylation level, especially that of CHH context, was decreased in apomictic cells, and the reduction in NEI cells was contributed primarily by that of retrotransposons. we suggest that the global reduction of DNA CHH methylation level in apomictic cells may derepress the retrotransposons, thus to alter expression of adjacent development regulatory genes, whereas DNA methylation on PCGs may positively regulate expression of critical biological processes, pathways and key genes to accomplish NPE initiation.