library_construction_protocol: Mouse diaphragm cell nuclei were isolated and purified following established protocols for scRNA-seq. Briefly, the fresh or frozen diaphragm tissue are minced and combined with 1ml of nuclear lysis buffer (NST) containing 0.1% NP40, 10mM Tris-HCl, 146mM NaCl, 1mM CaCl2, 21mM MgCl2, and 40 U/mL RNase inhibitor. The mixture is then incubated for 7 minutes to ensure complete nuclear lysis, which is confirmed by Trypan blue staining and microscopic examination. Next, 1ml of ST wash buffer (containing 10 mM Tris-HCl, 146 mM NaCl, 1 mM CaCl2, 21 mM MgCl2, 0.01% BSA, and 40 U/mL RNase inhibitor) is added to the lysed nuclei. The suspension is filtered through a 40 µm cell strainer, and the filtrate is collected in a 15mL centrifuge tube. The cell strainer is washed with additional ST wash buffer, and the wash solution is combined with the filtered nuclear suspension. The tube is then centrifuged at 500 g for 5 minutes at 4°C. The nuclear pellet is resuspended in 5 mL of PBS + 1% BSA, followed by centrifugation and resuspension in 100 µl of PBS + 1% BSA. Trypan blue staining is performed, and the cells are counted under a microscope The nuclei are counted, and the nuclear concentration is adjusted as needed by diluting with PBS + 1% BSA to achieve the optimal range for loading on the 10× Chromium chip. On-machine processing and cDNA library amplification are performed according to the instructions provided by the 10× Genomics Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 (1000268). For DNA library construction, the Chromium™ Single Cell 3'/5' Library Construction Kit (1000020) is used. The constructed libraries are sequenced using the PE150 sequencing mode on the Illumina Nova 6000 platform.