Bioscience Methods 2025, Vol.16, No.2, 83-99 http://bioscipublisher.com/index.php/bm 87 marker genes. For example, the satellite cell population highly expresses stem cell markers such as PAX7, the myoblast population highly expresses myogenic regulatory factors such as MYOD1 and MYOG, and the FAPs population is enriched in stromal cell markers such as PDGFRA. Single-cell analysis can also reveal some rare or previously under-characterized cell types. For example, a small number of cells with adipocyte characteristics were identified in scRNA-seq of skeletal muscle of mice and pigs, suggesting that FAPs in muscle may differentiate into adipocytes under certain conditions. Similarly, there may be low-abundance but functionally important cell populations in goat muscle tissue that can only be identified by single-cell sequencing. 3.2 Gene expression patterns in muscle lineages Single-cell transcriptomes can not only identify cell types, but also reveal the gene expression dynamics and regulatory networks of muscle lineage cells during differentiation. During muscle development, cells at different stages activate specific gene expression programs. By performing pseudotime analysis on single-cell data, the trajectory of cells from progenitor cells to differentiated cells can be reconstructed to observe when key genes are turned on or off. For example, in the trajectory of differentiation from myogenic precursors to myoblasts, cells in the early stage highly express PAX3/PAX7 and MYF5, representing the state of stem cells and primary progenitor cells; in the middle stage, factors that promote differentiation, such as MYOD1 and MEF2C, begin to be upregulated; and in the terminally differentiated myoblasts, muscle structural protein genes such as myosin heavy chain are significantly expressed (Zhou et al., 2022). Similar patterns are expected to be seen in single-cell data from goats. For example, assuming that along the trajectory from satellite cell activation to myoblast fusion, PAX7 will be highly expressed in the initial stage and decrease in the late stage, whereas MyoD/MyoG will be upregulated in the middle and late stages. This expression change has been confirmed in single-cell analyses of other species. At the same time, single-cell transcriptomes can also discover new genes or non-coding regulatory factors related to muscle development. Ye et al. (2023) found through transcriptome analysis that during the development of goat fetal muscle, the expression of structural-related genes such as integrin ITGA4/ITGA11 and troponin TNNT1/TNNT3 changed significantly, and participated in the regulation of myofilaments and adhesion plaques. The expression patterns of these genes revealed that they may play an important role in muscle formation. Although this study used bulk RNA-seq data, if single-cell sequencing is applied in the future, the expression of these genes can be further localized to specific cell subpopulations, thereby clarifying their role stages in the muscle lineage. In addition to coding genes, single-cell analysis also revealed differences in the expression of non-coding RNAs in different muscle cells. For example, Han et al. (2022) used second-generation sequencing to compare the expression of lncRNAs in skeletal muscle at different developmental stages of goats, identified a series of lncRNAs related to muscle growth, and predicted through co-expression analysis that they may regulate muscle structure or metabolic genes. At the single-cell level, if certain lncRNAs are highly expressed in satellite cells and lowly expressed in differentiated muscle fibers, it indicates that they may play a role in the maintenance or activation of muscle stem cells. Similarly, Shen et al. (2022) sequenced small RNAs in skeletal muscle of goats of different breeds and found that multiple miRNAs were closely related to muscle growth and intramuscular fat deposition. For example, miR-133, miR-378, etc. were upregulated during the period of vigorous muscle growth, and the regulatory targets of these miRNAs involved myocyte proliferation and adipose differentiation. Combining these findings with single-cell transcriptome data can verify the distribution of these miRNAs in specific cell types and further clarify their cell background of action. Recently, a study on Wu’an goats reported that miR-665 was highly expressed in the muscles of 1-month-old young goats, and could target and inhibit the apoptosis-related gene BCL2L11, thereby promoting myoblast proliferation (Feng et al., 2025). This result provides an example of miRNA regulating muscle cell fate and reflects the impact of miRNA expression changes in developmental periods (juvenile vs. young) on muscle growth. 3.3 Key findings from transcriptome data Using single-cell and other transcriptome data, researchers have made some important discoveries in skeletal muscle development in goats and related species. First, the regulatory mechanism of the balance between muscle and fat development has gradually become clear. Qiu et al. (2020) combined single-cell transcriptomics with
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