GSM9307277: CPUO_4; Homo sapiens; RNA-Seq
The human blood samples were centrifuged at 3,000 rpm for 15 minutes to separate plasma and cellular components. Plasma was aliquoted and stored at -80°C until further analysis. The cellular fraction was treated with RBC lysis buffer (Solarbio, China) to remove erythrocytes. The prepared human blood cells were washed with phosphate buffered saline (PBS) containing 0.04% bovine serum albumin (Sigma-Aldrich, USA). Cell viability was assessed by trypan blue staining with a threshold of >85%. Cell numbers were counted using an automated cell counter (Countess II FL, ThermoFisher) to achieve a final concentration of 700-1200 cells/μl. Single-cell suspension was added to the 10x Chromium chip according to the instructions for the 10X Genomics Chromium Single-Cell 3' kit (V3), with the expectation of capturing 8,000 cells. cDNA amplification and library construction were performed according to standard protocols. Libraries were sequenced by LC-Bio Technology (Hangzhou, China) on an IlluminaNovaSeq6000 sequencing system (double-end sequencing, 150 bp) at a minimum depth of 20,000 reads per cell.