METHODS FOR DETERMINING SPATIAL AND TEMPORAL GENE EXPRESSION DYNAMICS IN SINGLE CELLS

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Transcriptomes of individual neurons provide rich information about cell types and dynamic states. However, it is difficult to capture rare dynamic processes, such as adult neurogenesis, because isolation from dense adult tissue is challenging, and markers for each phase are limited. Here, Applicants developed Nuc-seq, Div-Seq, and Dronc-Seq. Div-seq combines Nuc-Seq, a scalable single nucleus RNA-Seq method, with EdU-mediated labeling of proliferating cells. Nuc-Seq can sensitively identify closely related cell types within the adult hippocampus. Div-Seq can track transcriptional dynamics of newborn neurons in an adult neurogenic region in the hippocampus. Dronc-Seq uses a microfluidic device to co-encapsulate individual nuclei in reverse emulsion aqueous droplets in an oil medium together with one uniquely barcoded mRNA-capture bead. Finally, Applicants found rare adult newborn GABAergic neurons in the spinal cord, a non-canonical neurogenic region. Taken together, Nuc-Seq, Div-Seq and Dronc-Seq allow for unbiased analysis of any complex tissue.