GSM9421943: Brain-5xFAD+/- Clec7a cKO; Mus musculus; RNA-Seq
Half brain samples were flash frozen on dry ice and sent to Azenta-GeneWiz for single-nuclei isolation. Single nuclei were isolated from tissue samples and nuclei suspensions were prepared under RNase-free conditions. Libraries were generated from single nuclei using a droplet-based single-cell/nucleus RNA-seq workflow that incorporates (i) reverse transcription of captured nuclear RNA, (ii) incorporation of nucleus-specific barcodes and unique molecular identifiers (UMIs) during cDNA synthesis, and (iii) PCR amplification of full-length cDNA followed by construction of Illumina-compatible sequencing libraries. Amplified cDNA was fragmented and end-repaired, A-tailed, and ligated to Illumina sequencing adapters and indexes. Final libraries were purified (SPRI bead–based cleanup) and assessed for yield and fragment size distribution (e.g., fluorometric quantification and capillary electrophoresis). Libraries were pooled at equimolar ratios and sequenced on an Illumina HiSeq 2000 platform using paired-end sequencing.