GSM7232715: Bone marrow, Stroke-treated, snATACseq, rep2; Mus musculus; ATAC-seq
Bone marrows were homogenized and filtered through 40 μm cell strainers to obtain single cell suspensions. Cell suspensions were were stained with a cocktail of antibodies against markers of the lymphoid lineage (CD3, CD4, CD8a, CD19 and Ter119) and mature neutrophils (Ly6G). Live, lymphoid lineage (CD3, CD4, CD8a, CD19 and Ter119) and neutrophil (Ly6G) negative BM cells were sorted using a FACS Aria II Fusion cell sorter (BD Biosciences, Inc.). Cells were centrifuged and incubated twice with Nuclei EZ Lysis Buffer on ice for 5 min each time. Lysed cells were washed and nuclei were incubated with 7AAD at 1g/ml for microscopic inspection of integrity before sorting on a FACS Aria II Fusion (BD Biosciences). Then, nuclei were further processed according to manufacturer's (10X) protocols at a concentration of 1000-5000 nuclei/µl. Single nuclei ATAC libraries were prepared using the 10x Chromium Next GEM Single Cell ATAC kit Libraries were performed according to the manufacter's instructions (single cell 3' v3.1 protocol and single cell 3' v3.1 with feature barcoding technology for Cell surface protein, 10x Genomics).