BioSample

Protocols: Mechanical separation of the cell types was performed at the developmental stage just before the onset of cell cleavage of reproductive cells; only this stage allows for clean separation of somatic and reproductive cells. Separation started with breaking the spheroids into parts in a Dounce homogenizer without harming the cells. Cell-size based separation of cell types was achieved by successive filtration on screens of different mesh sizes and a centrifugation step, just as previously described (Nematollahi et al., 2006). The clean separation of somatic cell sheets and reproductive cells was checked microscopically Cell-size based separation of cell types was achieved by successive filtration on screens of different mesh sizes and a centrifugation step Total RNA was extracted from separated cell types using the phenol-based TRI Reagent Total RNA isolated both from somatic and reproductive cells was prepared for RNA-seq analysis with three biological replicates each. For each cell type and biological replicate, 1 µg of total RNA was subjected to poly(A)+ RNA selection and mRNA fragmentation. This was followed by random primer cDNA synthesis and adapter ligation.