Sexual reproduction is indispensable for most eukaryotes. However, the definition of “sex” has remained controversial for centuries. Since the early 1930s, unisexual flowers have been used to investigate sex differentiation or determination in plants, but all unisexual flowers investigated to date result from prevention of sex cell differentiation in either the stamen or carpel, not link to the sex cells as originally proposed by Robbins and Pearson in 1933. To solve the problem, the concept of the “sexual reproduction cycle” (SRC) has been developed. The SRC represents a modified cell cycle that integrates changes of surrounding components, “sex” refers to heterogametes in the SRC (Fig. 17.1). Multicellular organisms are derived via the interpolation of multicellular structures (MCSs) into the two (diploid and haploid) intervals of SRC (Fig. 17.2). Therefore, sex differentiation occurs only in multicellular organisms and ensures heterogametogenesis. In plants, only differentiation of archegonia and antheridia fits the definition and could be considered as true sex differentiation, while differentiation of heterosporangia (e.g., stamens and ovules in angiosperms), is a form of pseudo sex differentiation. Finally, a third sex-related phenomenon, sexual behavior, has evolved in multicellular organisms to facilitate SRC completion. In plants, sexual behaviors include pollen tube growth, the attraction of pollinators to ensure that gametes meet, and the promotion of outcrossing. Unisexual flower development is one of the mechanisms of outcross promotion. The SRC-based comparison of plants with other eukaryotes provides an alternative reference framework for understanding the general principles of sex differentiation in all eukaryotes.