Project description:Spatial patterns of gene expression span many scales, and are shaped by both local (e.g. cell-cell interactions) and global (e.g. tissue, organ) context. However, most in situ methods for profiling gene expression either average local contexts or are restricted to limited fields of view. Here we introduce sci-Space, a scale-flexible method that retains single cell resolution while resolving spatial heterogeneity in gene expression at larger scales. As a proof-of-concept, we apply sci-Space to the developing mouse embryo (E14), capturing the approximate spatial coordinates of profiled cells from whole embryo serial sections.
Project description:We report the application of single-molecule-based sequencing technology for REST and its cofactors genome wide binding sites in E14 cells.We then combine these binding sirtes with REST regulating gene profiling, to understand REST binding and regulation in E14 cells.
Project description:Analysis of gene expression in Mouse E14-TG2a.IV strain ES cells Single end RNA-seq analysis of PolyA selected RNA from E14-TG2a.IV ES cells
Project description:More than 2x10E9 sequences made on Illumina platform derived from the genome of E14 embryonic stem cells cultured in our laboratory were used to build a database of about 2.7x10E6 single nucleotide variant. The database was validated using other two sequencing datasets from other laboratory and high overlap was observed. The identified variant are enriched on intergenic regions, but several thousands reside on gene exons and regulatory regions, such as promoters, enhancers, splicing site and untranslated regions of RNA, thus indicating high probability of an important functional impact on the molecular biology of this cells. We created a new E14 genome assembly including the new identified variants and used it to map reads from next generation sequencing data generated in our laboratory or in others on E14 cell line. We observed an increase in the number of mapped reads of about 5%. CpG dinucleotide showed the higher variation frequency, probably because of it could be target of DNA methylation. We performed a reduced representation bisulfite sequencing on E14 cell line to test our new genome assembly with respect to the mm9 genome reference. After mapping and methylation status calling, we obtained an increase of about 120,000 called CpG and we avoided about 20,000 wrong CpG calling.
Project description:We asked what genes are significantly differentially regulated in the spinal cord of SCI trkB.T1 WT and trkB.T1 KO mice. TrkB.T1 is upregulated shortly after SCI although the precise mechanisms underyling this upregulation are poorly understood. In the trkB.T1 null, we show less mechanical allodynia and better locomotor recovery following SCI. The microarray studies helped us to elucidate a signaling pathway that is differently regulated in the WT versus KO mice at 1 day after SCI. In this study, we did not examine gene changes within a genotype after SCI. Rather, we examined DGE by genotype at each time point. Spinal cord tissue from WT and KO mice in a sham condition (intact spinal cord) versus 1D, 3D and 7D following SCI was harvested for microarray analyses.