GSM8340279: Breast_adjacent_tissue_scRNAseq; Homo sapiens; RNA-Seq
Breast tissue was acquired from three Tru-Cut biopsies. The tissue was sectioned into small fragments and cryopreserved in FBS (Gibco) supplemented with 10% dimethyl sulfoxide (DMSO, Sigma). Upon thawing, the tissue was enzymatically dissociated using Accumax solution (Sigma) and incubated for 30 minutes. The resulting cell suspension was filtered through a 40 µm filter, followed by processing using the Dead Cell Removal Kit (Miltenyi Biotec #130090101) through two rounds to ensure sufficient viability. The scRNA-seq libraries were constructed using the Chromium Next GEM Single Cell 3´Reagent Kits v3.1 (Dual Index) following the manufacturer's instructions (10X Genomics). Briefly, the cell suspension was minimally manipulated as described above and loaded onto the Chromium Next GEM Chip G, avoiding bubble formation. Then the chip was inserted into the Chromium Single Cell Controller. Subsequently, the cDNA library was produced, cleaned, and amplified using 11 cycles of the specified PCR program. Libraries were constructed by adding the appropriate adapters and indices (Dual Index TT Set A) provided by the manufacturer. The final amplification was carried out with 12 cycles of the specified PCR program.