The early 1900s delivered many foundational discoveries in genetics, including re-discovery of Mendel's research and the chromosomal theory of inheritance. Following these insights, many focused their research on whether the development of separate sexes had a chromosomal basis or if instead it was caused by environmental factors. It is Dr Nettie M. Stevens'Studies in spermatogenesis(1905) that provided the unequivocal evidence that the inheritance of the Y chromosome initiated male development in mealworms. This result established that sex is indeed a Mendelian trait with a genetic basis and that the sex chromosomes play a critical role. In Part II ofStudies in spermatogenesis(1906), an XY pair was identified in dozens of additional species, further validating the function of sex chromosomes. Since this formative work, a wealth of studies in animals and plants have examined the genetic basis of sex. The goal of this review is to shine a light again on Stevens’Studies in spermatogenesisand the lasting impact of this work. We additionally focus on key findings in plant systems over the last century and open questions that are best answered, as in Stevens' work, by synthesizing across many systems.This article is part of the theme issue ‘Sex determination and sex chromosome evolution in land plants’.