GSM8146176: mouse DRG cell culture, 14 days post infection with Neurog2-Neurog1-DsRed, rep1; Mus musculus; RNA-Seq
Primary DRG cells infected with CAG-Neurog2-Neurog1-DsRed were harvested at 14 dpi in an FCS-coated Falcon. Cells were filtered through a 100 µm cell filter (pluriSelect, Cat#43-10100-40) and sorted using a BD FACS Aria III Cell Sorter (BD Biosciences) based on DsRed fluorescence. Cells were collected through a 100 µm nozzle into 0.04% BSA in 1xPBS. Cells were centrifuged and diluted to a concentration of 1,000 cells/µl in 0.04% BSA in 1xPBS prior to single cell RNA sequencing. Single-cell isolation and library preparation was performed via 10x Genomics Chromium Next GEM 3' kit v3.1 following the manufacturer's instruction. In brief, FACS-sorted cells were subsequently loaded on a 10x Genomics Chip G and encapsulated into Gel Bead-in-Emulsion (GEM) droplets, each containing a single cell along with a unique barcoded bead. Within these GEMs, the mRNA from each cell was reverse transcribed into complementary DNA (cDNA) using reverse transcriptase, with the process occurring on the uniquely barcoded beads. The resulting cDNA was then recovered from the GEM droplets and amplified to prepare sequencing libraries using PCR. During library preparation, additional sequences necessary for sequencing, such as adapter sequences, were incorporated into the cDNA fragments.