Association of morphological markers of flower buds and anthers with the pollen developmental stage in Vicia faba L. (Fabaceae).

The precise identification of microspore and pollen at the optimal developmental stages to be induced towards embryogenesis (vacuolated microspores and young pollen) is essential for induction of in vitro androgenesis in plants. Such identification is not always easy, and it is especially difficult in recalcitrant species such as Vicia faba. The present study evaluates the relationship between floral bud and anther morphometric parameters, and microspore/pollen developmental stages in V. faba using various methods including fluorescent staining, differential interference contrast microscopy, and morphometry. We measured flower bud and anther length and width, grouping them at different intervals, and performed a detailed microscopical and anatomical analysis of buds, anthers and microspores/pollen at different stages. Our results demonstrated that flower buds in V. faba exhibit complex and irregular morphologies, with considerable variation in both sepal length and shape. Furthermore, the determination of microspore and pollen developmental stages in this species is constrained by pronounced developmental asynchrony and strong genotype dependence. Although anther length measurements correlate closely with microspore and pollen developmental stages, their practical use can be challenging. Therefore, measuring flower bud length, while excluding sepals, remains the most practical criterion for routine applications. Combining this refined morphometric approach with microscopic validation appears to be the most effective strategy for improving the identification of flower buds containing microspores or pollen at developmental stages suitable for androgenesis induction in this recalcitrant legume species.