© 2023, Korean Society for Plant Biotechnology.Cycloastragenol (CAG) is a compound isolated from Astragalus membranaceus, which significantly stimulates telomerase activity and cell proliferation and prevents some diseases in humans. Plants endure different stress conditions leading to physiological, biochemical, and molecular perturbations affecting their growth and development. CAG could prevent and remedy these disorders in plants. A high-throughput sequencing methodology was carried out to determine how CAG affects different signaling mechanisms in plants and to overview the post-transcriptional CAG responses analysis. A small RNA sequencing strategy was employed in A. thaliana roots calli under CAG treatment to assess known microRNAs and evaluate their eventual functions in the affected pathways. The data revealed 59 differentially expressed known miRNAs targeting 910 genes. The results show that several miRNAs are associated with metabolic and stress tolerance signaling pathways, including miR159b-5p and its target genes, which have an essential transducing role in different environmental stresses, and miR3434, which is involved in root growth flowering and stress prevention.