Expression profiling by high throughput sequencing
Summary
Here, we present a comprehensive transcriptome roadmap of the critical developmental time window prior to gastrulation by sequencing and analysis of 1724 individual cells from 28 pregastrula mouse embryos. Our analyses reveal the cellular substructure in EPI through clustering cells into anterior, transition and posterior states. The induction of the primitive streak (PS) is achieved via a transition state of a high cellular variability and up-regulation of the PS signature genes. Moreover, the evolvement of these three cellular states corresponds well with the transition from mouse naïve embryonic stem cells (i.e mESCs) to primed epiblast stem cells (i.e. mEpiSCs). In addition, dissect the substructure of visceral endoderm (VE) and identify novel markers of anterior VE (AVE). Importantly, we also characterize the lineage-specific dynamics of X-chromosome inactivation (XCI), demonstrating that reversal of XCI (i.e. X reactivation) in the EPI lineage occurs before imprinted silencing of the paternal X-chromosome is completely established, and the ensuing random XCI is highly asynchronous. VE displays faster progression of imprinted XCI compared to the extra-embryonic ectoderm (ExE). In summary, our data not only provide new insight into the regulation of lineage specification and XCI dynamics during the early embryogenesis but also shed light on the in vivo counterparts of stem cells including hypothesized formative pluripotency.
Overall design
In total, 1724 cells with three types of mice crossing were utilized. 21 blastocyst cells used for X-chr interactivation analyses.
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