GSM9745925: hippocampal tissues of AAV2/9-Syn-cre injected Bcas2F/F mice; Mus musculus; RNA-Seq
Tissue was added to 1 ml of pre-chilled lysate, sheared into small pieces of approximately 1 mm, 2 ml of pre-chilled lysate was added, transferred to a tissue homogenizer (885300-0007; Kimble Dounce Homogenizer ), and ground slowly 5 times. The tissue was incubated on ice for 5 min, followed by the addition of 4 ml of pre-cooled 2% BSA dilution, and the reaction was terminated by pipetting with a barrel pipette. Samples were passed through a 30um cell sieve, and the filtrate was collected and centrifuged at 300 g for 5 min at 4°C. The supernatant was discarded. The cells were resuspended with 5 ml of 2% BSA, centrifuged at 4°C for 5 min at 300 g. The supernatant was discarded and the cells were resuspended with 500 ul of resuspension solution (1xPBS, 2% BSA, 0.1% RNase inhibitor). Stain with PI (Propidium Iodide) and sort to remove debris impurities using a BD FACSAria II flow cytometer. Cells were collected and counted. Single-cell nuclear 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 Illumina NovaSeq 6000 sequencing system (double-end sequencing, 150 bp) at a minimum depth of 20,000 reads per cell.