GSM9316823: sublib3; Mus musculus; RNA-Seq
Animals were deeply anesthetized with ketamine–xylazine and perfused transcardially with 10 mL PBS. Brains were collected, and brain single-cell suspensions were prepared using the Adult Mouse Brain Dissociation Kit (Miltenyi) according to the manufacturer's instructions. CD45+ cells were then enriched by staining with CD45 magnetic microbeads and applying them to an LS column (Miltenyi) following the manufacturer's protocol. Isolated cells were pelleted and fixed overnight in 10x Genomics Fixation Buffer (10x Genomics). Samples from 4–5 mice per experimental time point were pooled before submission to the University of Illinois Urbana-Champaign Genomics Core for processing with the 10x Genomics Chromium Fixed RNA Profiling (Flex) Gene Expression assay (10x Genomics). All libraries were prepared and sequenced together on the Illumina NovaSeq X Plus System, generating 28 × 90 nt reads. Notably, probes targeting the SARS-CoV-2 ORF1ab and nucleocapsid (N) genes were custom-designed and spiked into the Chromium Mouse Transcriptome Probe Set (v1.0.1), following the manufacturer's instructions. Probe sequences were as follows: ORF1ab: TACTAGTGCCTGTGCCGCACGGTGTAAGACGGGCTGCACTTACACCGCAA; ATAGATTACCAGAAGCAGCGTGCATAGCAGGGTCAGCAGCATACACAAGT; GAGCAAGAACAAGTGAGGCCATAATTCTAAGCATGTTAGGCATGGCTCTA. N: CCATTCTGGTTACTGCCAGTTGAATCTGAGGGTCCACCAAACGTAATGCG; AGTTGTAGCACGATTGCAGCATTGTTAGCAGGATTGCGGGTGCCAATGTG; TGGAACGCCTTGTCCTCGAGGGAATTTAAGGTCTTCCTTGCCATGTTGAG.