GSM8886759: Sample N2; Mus musculus; RNA-Seq
The femur or cranium tissues of Ctsk-Cre; Nfatc1-Dre; LGRT neonatal mice were dissected and cut into pieces. Then, fibrous callus was digested in 1 ml pre-warmed digestion solution (3 mg/ml collagenase I (Sigma, USA) in αMEM) for 45 min in a water bath at 37 ℃ with gentle agitation every 10 min. Then, the mixture of digested minced bones and target cells were treated with red blood cell lysate on ice for 3 min. The cells were then isolated using Fluorescence Activated Cell Sorting (FACS) , sorting the femur and cranium into ZsGreen+ tdTomato-, ZsGreen- tdTomato+, and ZsGreen+ tdTomato+ cells, which were ultimately analyzed using scRNA-seq. cDNA was purified using DynaBeads® MyOne™ Silane Beads (Life Technologies, 37002D). The concentration of cDNA samples was quantified using the Qubit dsDNA Assay Kit (Life Technologies, Q328520), and high-sensitivity DNA analysis was performed using the Agilent High Sensitivity DNA Kit (Agilent, 5067-4626). DNA fragments were selected and purified using the SPRIselect Reagent Kit (Life Technologies, B23318), ultimately obtaining high-quality single-cell gene expression data.