This study delves into the intricate cellular landscape of the ovary, aiming to unravel the complexities of its various cell populations and their dynamics during the estrus cycle leveraging single-cell RNA sequencing. This dissertation achieves two primary objectives: first, identifying stromal cell subclusters and localization of novel collagens within the bovine ovary, and second, identifying unique cell types in the ovarian cortex and medulla while tracking their changes across different estrus cycle stages and probing into the origins of theca cells within the ovarian stroma. Major findings include the discovery of putative distinct cell types, such as theca interstitial cells and migratory stromal cells, along with shifts in cell proportions from the follicular to luteal phases. Notably, the presence of the hedgehog signaling pathway within the cortex confirms previous reports in cattle and other species and suggests a potential role in the follicular recruitment of theca cells. Furthermore, the investigation extends to the medullary region, uncovering diverse cell populations encompassing endothelial cells, smooth muscle cells, immune cells, neuronal cells, and stromal cells. The luteal phase emerges as a period marked by intense proliferation of endothelial cells, vital for angiogenesis and the sustained function of the corpus luteum. While this research enhances our comprehension of the ovarian microcosm, it also underscores the necessity of further functional exploration. Future endeavors will focus on delving deeper into the functional implications of these findings in the realm of reproductive biology and the advancement of assisted reproductive technologies.