Project description:Cells continuously communicate with the neighboring cells during development. Direct interaction of different cell types can induce molecular signals dictating lineage specification and cell fate decisions. The current single-cell RNAseq (scRNAseq) technology cannot study cell contact dependent (or niche specific) gene expression due to the loss of spatial information. To overcome this issue and determine cell contact specific gene expression during embryogenesis, we performed RNA sequencing of physically interacting cells (PICseq) and assessed alongside our single cell transcriptomes (scRNAseq) derived from developing mouse embryos between embryonic day (E) 7.5 and E9.5. Analysis of PICseq data identifies an interesting suite of gene expression signatures depending on neighboring cell types. For instance, neural progenitor (NP) cells expressed Nkx2-1 when interacting with definitive endoderm (DE) and DE cells expressed Gsc when interacting with NP. Based on the identified cell contact specific genes, we devised a means to predict the neighboring cell types from individual cell transcriptome. We further developed spatial-tSNE to show the pseudo-spatial distribution of cells in a 2-dimensional space. In sum, we suggest an approach to study niche specific gene regulation during embryogenesis.
Project description:Primordial germ cell mRNA profiles from cells microdissected from e6.5, e7.5 and e8.5 embryos, e7.5 somatic neighbours and Blimp1-KO mice were generated by single cell library construction and sequencing in duplicate using Applied Biosystems SOLiD sequencer. Single cell library construction is described in: Tang f. et. al, Nature Protocols (2010), Vol. 5, p.516.
Project description:LV-EF1A-H2B-tdTomato-30N barcode virus was injected into the amniotic cavity (AC) of embryos at E7.5, using ultrasound guided in-utero nano-injections. E9.5 and E10.5 whole embryos were collected, dissociated, and sorted for tdTomato. Sorted cells were subjected to single cell RNA sequenced using 10X Genomics sequencing.
Project description:Understanding processes how the early stage kidney precursor gives rise to metanephric mesenchyme, which is a committed progenitor cells of adult kidney is important for the regeneration of kidney in vitro. The combination of fluorescent activated cell sorting (FACS) plus microarray analysis offers a powerful, efficient and effective method for the creation of global gene expression profiles of the developing kidney precursors. Those gene expression data provides insights into not only the stage specific marker genes but also the signals working in each population, which should be informative for the directed differentiation of pluripotent stem cells in vitro. Osr1-GFP knock-in mice were used to isolate kidney precursor cells from embryos at E8.5, E9.5 and E11.5. At E9.5 and E11.5 embryos, to identify the differences between nephron progenitors and surrounding mesenchyme, nephron progenitor populations were further enriched by gating Osr1-GFP positive Integrin alpha8 positive Pdgfr alpha negative population and compared with Osr1-GFP positive cells other than that gate. RNA was isolated from cells and the gene expression profiles were determined by microarrays.
Project description:We analyzed wildtype and miR-302 knockout embryos at E7.5 and sorted neural crest using Wnt1-Cre at E8.5 and Sox9 at E9.5 to capture miRNA differences during neural crest development
Project description:We analyzed wildtype and miR-302 knockout embryos at E7.5 and sorted neural crest using Wnt1-Cre at E8.5 and Sox9 at E9.5 to capture transcriptomic differences during neural crest development
Project description:Transcription profiling by high throughput sequencing of primordial germ cells from e6.5, e7.5, e8.5 embryos and e7.5 somatic neighbours and Blimp1-KO mice
Project description:The placenta is a transient organ that is repsonsible for multiple processes during pregnancy. Here we aim to study the development of the placenta using mouse models at embryonic day (e) e7.5, e8.5 and e9.5 by integrating RNA-seq data at the three timepoints to identify important genes, inferring their interaction networks, and predict novel regulators of placental development.
Project description:UHRF1 is a chromatin-binding protein essential for maintaining DNA methylation and histone modification states, yet its integrated role in vivo remains incompletely understood. To define its function, we generated conditional Uhrf1 knockout embryonic stem cells (ESCs) and embryos. Uhrf1⁻/⁻ ESCs exhibited near-complete loss of 5mC and 5hmC but maintained pluripotency, whereas Uhrf1-null embryos developed normally until E8.5 and then failed to further develop by E9.5.