Molecular diversity of olfactory bulb projection neurons [single-cell RNA-seq]

To characterize the molecular diversity of olfactory bulb projection neurons we used viral targeting and Fluorescence Activated Nuclei Sorting (FANS) to enrich for olfactory bulb projection neurons, and single-nuclei RNA sequencing (sn-RNA seq) to comprehensively characterize their transcriptomes. To isolate GFP-labelled nuclei, 3 individual replicates of AON and PCx-injected mice were used. Ipsilateral and controlateral sides were minced separately and placed into two different tubes. The minced tissue was gently homogenized in Nuclei PURE Lysis Buffer and 10% Triton X-100 using an ice-cold dounce and pestle, and filtered two times through a 40 μm cell strainer on ice. After centrifuging at 500 rpm for 5 min at 4 °C, the supernatant was aspirated and gently resuspended in 500 μl of cold buffer (1x of cold Hanks' Balanced Salt Solution HBSS, 1% nuclease-free BSA, RNasin Plus and 1/2000 DRAQ5). Our study identifies molecularly distinct subtypes of mitral and tufted cells. we used AAV viral targeting and Fluorescence Activated Nuclei Sorting (FANS) to enrich for olfactory bulb projection neurons, and single-nuclei RNA sequencing (10x Genomics)