title: GSM9462553: wks28 scRNA-seq from mandible; Mus musculus; RNA-Seq
library_construction_protocol: Mandibles were dissected from sixteen 8-week-old male C57BL/6 mice (GemPharmatech Co., Ltd.). Soft tissues, molars, incisors, and bone posterior to the condyle were removed. The isolated bone tissue was enzymatically digested in 3 mg/mL collagenase type I (Gibco) and 4 mg/mL dispase II (MilliporeSigma) for 60 minutes at 37°C. Resuspend the single cells in MACS buffer (0.5% BSA, 2 mM EDTA) and add CD45 MicroBeads (Miltenyi). Incubate the mixture for 15 minutes in a 4°C refrigerator. Following incubation, wash the cells with MACS buffer. Then, apply the cell suspension onto the MACS LS column placed within the separator's magnetic field. Collect the flow-through fraction which were CD45⁻ cells. Perform a second magnetic separation step on the collected CD45⁻ cells using anti-Ter119 MicroBeads (Miltenyi). After generate CD45-/Ter119- single-cell suspensions, 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 Illumina NovaSeq 6000 sequencing system (double-end sequencing, 150 bp) at a minimum depth of 20,000 reads per cell.