GSM8488404: S7d, Library 1; Mus musculus; RNA-Seq
Mice were anesthetized with ketamine and xylazine, and transcardially perfused with PBS. Brains were immediately harvested and placed in the dissection solution (0.8% glucose and 15 mM HEPES in HBSS) on ice. The lateral SVZ from the hemisphere ipsilateral to the cortical PT site was microdissected according to published protocols53. The SVZ tissue was digested with papain (Worthington, 1200 units per 5 mice) in PIPES solution [120 mM NaCl, 5 mM KCl, 50 mM PIPES (Sigma-Aldrich), 0.6% glucose, 1× Pen/Strep in water, pH adjusted to 7.6] for 10 min at 37°C, and mechanically dissociated into single cells with fire-polished glass Pasteur pipettes after adding ovomucoid (Worthington, 0.7 mg per 5 mice) and DNase I (Invitrogen,  1,000 units per 5 mice) as described54. Myelin debris was removed by centrifuging in 22% Percoll (Sigma-Aldrich) for 10 min at 4ºC without brakes. Single-cell RNA sequencing was performed using the mCEL-Seq2 protocol, an automated and miniaturized version of CEL-Seq2 on a mosquito nanoliter-scale liquid-handling robot. Eight libraries with 192 cells each were synthesized.