Development of multicellular pollen of Eriobotrya japonica Lindl. through anther culture

Eriobotrya japonica Lindl. is a worldwide known tree important for its use as horticultural and ornamental plant, especially in sub-tropical and Mediterranean countries. Microspore embryogenesis through in vitro anther culture is a widely used method to generate genetic variability by obtaining gametic or somatic embryos with many applications for plant breeding. In this work, a protocol has been set up for anther culture in loquat which resulted in the formation of multicellular pollen as a first step to further attempt haploid–plant production via pollen-derived structures. The response of nine of the most widely grown loquat cultivars to anther culture has been evaluated, and four cultivars being selected due to their higher response. The occurrence of anther swelling and the development of calli were analyzed as typical morphological features and potential markers that accompany pollen induction and reprogramming in many systems. Microscopical analysis in responsive anthers and the comparison with the normal gametophytic pollen development was carried out to characterize the cellular changes promoted by the treatment in the anther. The presence of multicellular pollen in the in vitro system developed here indicated the switch of developmental programme which constitutes a crucial step in the design of protocols for the regeneration of microspore-derived embryos and plants. Monitoring structural analysis at different times of the culture revealed specific features of the early microspore embryogenic pathway as well as the cellular organization changes.