Vitamin D, an essential steroid hormone in the human body, plays an important role in not only the regulation of calcium and phosphorus metabolism, but also in various physiological processes, such as cell differentiation and apoptosis, inflammation, and insulin resistance. Vitamin D receptors are widely distributed in male and female reproductive systems, suggesting that vitamin D is essential for fertility. Because vitamin D deficiency is highly prevalent around the world, this review aims to discuss the potential functions of vitamin D in male and female reproductive systems and the associations between vitamin D and assisted reproductive technology (ART) outcomes. Vitamin D is involved in many physiological reproductive processes, including steroidogenesis, spermatogenesis, and acrosome reaction. It is correlated with sperm quality, ovarian reserve, polycystic ovarian syndrome, and endometriosis, among others. Controversial clinical findings on vitamin D levels and ART outcomes were revealed in this review, and demonstrations of efficacy for human fertility in randomized controlled trials of vitamin D supplementation are notably lacking. Thus, further studies are highly required involving molecular mechanisms among different species and human populations, as well as randomized controlled trials.