The myostatin (MSTN) gene is of interest in the livestock industry because mutations in this gene are closely related to growth performance and muscle differentiation. Thus, in this study, we established MSTN knockout (KO) quail myoblasts (QM7) and investigated the regulatory pathway of the myogenic differentiation process. We used clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 to generate MSTN KO QM7 cells and subsequently isolated a single cell-derived MSTN KO QM7 subline with 10- and 16-nucleotide deletions that induced translational frameshift mutations. The differentiation capacity and proliferation rate of MSTN KO QM7 cells were enhanced. We conducted next-generation-sequencing (NGS) analysis to compare the global gene expression profiles of wild-type (WT) QM7 and MSTN KO QM7 cells. Intriguingly, NGS expression profiles showed different expression patterns of p21 and p53 in MSTN KO QM7 cells. Moreover, we identified downregulated expression patterns of leukemia inhibitory factor and DNA Damage Inducible Transcript 4, which are genes in the p53 signaling pathway. Using quantitative RT-PCR (qRT-PCR) analysis and western blotting, we concluded that p53-related genes promote the cell cycle by upregulating p21 and enhancing muscle differentiation in MSTN KO QM7 cells. These results could be applied to improve economic traits in commercial poultry by regulating MSTN-related networks.

肌肉生长抑制素（MSTN）基因在畜牧业中很受关注，因为该基因的突变与生长性能和肌肉分化密切相关。因此，在这项研究中，我们建立了MSTN基因敲除（KO）鹌鹑成肌细胞（QM7），并研究了成肌分化过程的调控途径。我们使用成簇的规则间隔的短回文重复序列（CRISPR）/ Cas9生成MSTN KO QM7细胞，随后分离出具有10和16个核苷酸缺失的单细胞衍生MSTN KO QM7亚系，该亚核苷酸引起翻译移码突变。MSTN的分化能力和增殖率KO QM7细胞得到增强。我们进行了下一代测序（NGS）分析，以比较野生型（WT）QM7和MSTN KO QM7细胞的全局基因表达谱。有趣的是，NGS表达谱显示了MSTN KO QM7细胞中p21和p53的不同表达模式。此外，我们确定了白血病抑制因子和DNA损伤诱导转录本4的下调表达模式，它们是p53信号通路中的基因。使用定量RT-PCR（qRT-PCR）分析和蛋白质印迹，我们得出结论，p53相关基因通过上调p21和增强MSTN的肌肉分化来促进细胞周期KO QM7细胞。这些结果可用于通过调节与MSTN相关的网络来改善商业家禽的经济特性。