The sub-ventricular zone was isolated and dissociated as described above for “Single cell RNA-seq using the 10x Genomics Chromium single cell technology”. In some cases, mice were given a 1mg/mouse dose of 5-ethynyl-2’-deoxyuridine (EdU) via intraperitoneal injection 4 hours prior to euthanasia. Antibody staining was carried out in FACS buffer at the following dilutions: PROM1-Biotin (eBioscience Cat.#13–1331-80 [1:300]), EGF-AlexaFluor 647 (Life Technologies Cat. #E35351 [1:300]), CD24-PacBlue (eBioscience Cat.#48–0242-80 [1:400]), CD31-PE (eBioscience Cat.#12–0311-81 [1:50]), CD45-BV605 (Biolegend Cat.#103139 [1:100]), Strep-PECy7 (eBioscience Cat.#25–4517-82 [1:500]), O4-PE (Miltenyi Cat.#130–099-211 [1:50]), CD317-FITC (Biolegend Cat.#127002 Clone:927 [1:50]) for one hour at 4°C. Samples were washed with 5mL of FACS buffer. Secondary staining was performed using Strep-PECy7 (eBioscience Cat.#25–4517-82 [1:500]) in FACS buffer at 4°C. Samples were washed with 5mL of FACS buffer, and resuspended in media containing 1μg/mL propidium iodide (Biolegend). Fluorescent-minus-one controls were used to set positive gates in each experiment. Cell populations were defined as follows:
BST2-positive and BST2-negative aNSCs/NPCs were subjected to subsequent analyses including bulk RNA-sequencing, EdU cycle analysis, and STAT1 staining.
BST2-positive and BST2-negative aNSCs/NPCs were subjected to subsequent analyses including bulk RNA-sequencing, EdU cycle analysis, and STAT1 staining.