【Objective】 As a ruminant animal, cattle has a long muscle growth and development cycle. However, the regulatory mechanism of chromatin accessibility on bovine longissimus dorsi is not clear during the development of longissimus dorsi. This study aimed to explore the differential chromatin accessibility of the longissimus dorsi muscle of Angus cattle and Simmental cattle, screen the key transcription factors affecting bovine muscle growth, and then construct the TF-gene regulatory network, so as to provides the theoretical basis and new molecular target for further study of the epigenetic regulation mechanism of bovine muscle development.【Method】 The longissimus dorsi muscle tissues of 24-month-old Angus cattle (AGS) and Simmental cattle (XM) with the same feeding and management methods were collected, and the amino acid content was detected according to the national standard Determination of amino acids in food (GB 5009.124-2016). SPSS 9.4 software was used to analyze the significant difference of amino acid content in longissimus dorsi muscle between groups by t test. The collected longissimus dorsi muscle tissue was subjected to ATAC-seq sequencing. By identifying the chromatin open area (peak), peak analysis, GO/KEGG functional enrichment analysis, and transcription factor binding site (motif) enrichment analysis were performed. Combined with the differentially expressed genes (DEGs) obtained from the previous RNA-seq sequencing, a transcription factor-gene (TF-gene) regulatory network was constructed based on the GTRD database and the OmicShare Tools.【Result】 The results showed as follows：(1) The content of lysine (Lys), serine (Ser), arginine (Arg), histidine (His) and glutamic acid (Glu) in longissimus dorsi muscle of AGS group were significantly higher than those of XM group (P < 0.05); the content of proline (Pro) was the opposite (P < 0.05), indicating that the amino acid metabolic phenotype was significantly different between the two groups. (2) Chromatin accessibility was generally conserved between the two groups. A total of 29 140 peaks were detected in Angus cattle and 28 781 peaks were detected in Simmental cattle. The chromatin open regions between the two groups were mainly distributed at the transcription start site (TSS) ± 2 kb, and accounted for more than 84.57 % in the intron region, distal intergenic region and promoter region. (3) A total of 6 185 differential peaks were identified between the two groups, of which 5 030 peaks were up-regulated and 1155 were down-regulated. After the differential peak were annotated to related genes, GO/KEGG enrichment analysis showed that the differential peak-related genes were mainly enriched in classical muscle development-related pathways, such as anatomical morphogenesis, muscle structure development, actin filament-mediated, Hippo signaling pathway, MAPK signaling pathway, calcium signaling pathway, and actin cytoskeleton regulation. (4) Motif enrichment analysis showed that among the top 20 transcription factor binding motifs, the top four belonged to the MEF2 transcription family, which were MEF2C, MEF2A, MEF2D, and MEF2B, respectively. Further focusing on the intersection analysis of AGS-specific peak and differentially expressed genes obtained by previous RNA-seq sequencing, it was found that the binding sites of MEF2B and MEF2D were the most enriched, and the TF-gene regulatory network with MEF2B and MEF2D as the core transcription factors was successfully constructed. A total of 13 target genes involved in muscle development regulation were screened, including ACTA1, CKM, CLCN1, SLN, and MYOZ3. IGV visualization confirmed the presence of MEF2B and MEF2D binding motifs in the promoter region of the above target genes, and highly overlapped with the open region of ATAC-seq.【Conclusion】 In this study, the combined analysis of ATAC-seq and RNA-seq data revealed the differences in chromatin accessibility between Angus cattle and Simmental cattle during the development of longissimus dorsi muscle, and screened MEF2B and MEF2D as key transcription factors (TFs) regulating bovine muscle growth. The TF-gene regulatory network with MEF2B and MEF2D as core transcription factors was constructed, which provided a theoretical basis for further analysis of the epigenetic mechanism of muscle development in beef cattle and functional verification of CRISPR-Cas9.