title: GSM8504709: Veh_3; Mus musculus; RNA-Seq
library_construction_protocol: After decapitation, the brain was removed, and the DVCs were isolated from the brainstem under a surgical microscope. A 1 mm coronal section of the hindbrain from bregma -7.0 to -8.0 mm was cut with a razor blade. From the coronal section, a 1.4 mm square containing the DVC was snap-frozen. Samples from the same experimental condition were pooled (n = 5 mice per sample) and nuclei were extracted. Sorting of 2n nuclei was performed by flow cytometry using a BD FACSAria IIIu Influx cell sorter (BD Biosciences). The gating was set according to size and granularity using FSC and SSC to capture singlets and remove debris. To detect DraQ5-positive nuclei fluorescence was set at 647 nm and 670 nm. Each sample was sorted into separate tubes, each with a total of 20,000 nuclei per 40 µl. Sequencing libraries were generated using 10x Genomics Chromium Single-Cell 3′ Reagent kit according to the standardized protocol. Paired-end sequencing was performed using an Illumina NovaSeq 6000.