GSM9470574: P7 SVZ multiomic data, snRNA-seq; Mus musculus; RNA-Seq
For single-cell isolation of the embryonic (E14, E17) cortex, the meninges were removed and tissue immediately dorsal to the lateral ventricles—including the cortical layers and the V-SVZ—was collected. For single-cell isolation of the postnatal (P1, P2, P6, P7) V-SVZ, the cortex was removed to expose the lateral ventricles, and the dorsolateral V-SVZ was microdissected out and collected. During dissections, collected tissue was placed in either DMEM (Gibco) or Hibernate A without calcium (BrainBits) supplemented with 2% B27 (Life Technologies) and 500 µM L-glutamine (Cedarlane) and left on ice until subsequent processing steps. For some datasets (E17 cortex multiomic, P2 V-SVZ n2 multiomic, P7 V-SVZ multiomic), tissue collection was immediately followed by an enzymatic digestion step. Samples were incubated in 20 units/mL Papain with 1 mM L-cysteine, 0.5 mM EDTA, and 0.005% DNase I (Worthington) dissolved in Hibernate A medium without calcium (BrainBits) and DRAQ5 (Abcam) diluted 1:3000 for 30 minutes at 37°C to simultaneously enzymatically digest the tissue and label nuclei. Following enzyme treatment, cells were spun down at 300 g for 5 minutes and resuspended in media, either DMEM (Gibco) or Hibernate A medium without calcium, supplemented with 2% B27 (Life Technologies) and 500 µM L-glutamine (Cedarlane). Following enzymatic digestion or immediately following collection, tissue was triturated using blunt needles with sequentially decreasing diameters (McMaster-Carr). Cells were then spun down at 300 g for 5 minutes and resuspended in 0.25% BSA (Jackson) in HBSS, after which the cell suspension was filtered through a 40 µm cell strainer and incubated in 1:1000 propidium iodide (PI) (Abcam) to label dead/damaged cells. Samples were then FACS sorted using a MoFlo Cell Sorter (Beckman Coulter) or Aria Fusion A (Becton Dickinson) to enrich for viable (PI-negative, DRAQ5-positive) cells, which were collected in media, either DMEM or Hibernate A without calcium, supplemented with 2% B27 and 500 µM L-glutamine. Libraries were constructed according to the manufacturer's instructions (Single Cell Multiome ATAC + Gene Expression v1, 10x Genomics).