GSM8768489: FRDA brain cells did not receive mitochondria; Mus musculus; RNA-Seq
Cells were isolated from three mice and pooled for scRNA-seq Brain tissue dissociation was performed using a modified protocol from the Adult Brain Dissociation Kit (Miltenyi Biotec 130-107-677). Mice were deeply anesthetized with a Ketamine/Xylazine mixture and underwent rapid decapitation to minimize brain tissue damage. The extracted brains were washed three times with ice-cold Dulbecco's phosphate-buffered saline (D-PBS) containing calcium, magnesium, glucose, and pyruvate (Thermo Fisher Scientific 14287080). Tissue pieces were collected by centrifugation at 400 g for 5 minutes and dissociated with enzyme mixes 1 and 2 in gentleMACS C Tubes (Miltenyi Biotec 130-093-237) for 30 minutes using the gentleMACS Octo Dissociator with Heaters (Miltenyi Biotec 130-096-427), according to the manufacturer's instructions. Digested samples were quenched with ice-cold D-PBS and filtered through a 70 μm cell strainer. After debris and red blood cell removal, following the manufacturer's instructions, the cell pellet was washed with FACs-BL and stored at 4°C for further processing. Cells were processed using the Chromium Controller and the Chromium Next GEM Single Cell 3' Reagent Kits (10x Genomics). The libraries were sequenced via Illumina Novaseq 6000 sequencer.