Examining the relationship between bud, anther morphology and developmental stages of the male gametophyte in onion (Allium cepa L.)

Abstract

Correlations between the morphological characteristics of flower buds and the developmental stages of the male gametophyte have practical implications as a reliable indicator for selecting suitable plant material for androgenesis. However, the application of androgenesis in onions has been challenging, despite extensive research on anther culture. This research paper presents findings on the associations between flower bud length and diameter, anther length and diameter, and the developmental stages of the male gametophyte across ten onion genotypes. All the morphological parameters studied in this study were positively correlated with stages of microspore development. The most responsive stages for anther culture in most of the crops are the late uninucleate and early binucleate stages. Although the bud stage and microspore development stage among the genotypes varied but this stage was typically observed in flower buds with a length of 2.5 to 3.5 mm and a diameter of 2.0 to 2.7 mm. Likewise, the late uninucleate and early binucleate stages were found in anthers with length of 1.2 to 1.6 mm and diameter of 0.55 to 0.75 mm. Additionally, we examined the utility of the aforementioned morphological parameters in identifying microspore developmental stages. Based on our results, we propose a combination of flower bud length, diameter, and the green colour intensity of anthers as easily applicable, rapid, and accurate criteria for identifying microspore developmental stages. Different methods were employed to evaluate the efficiency of DAPI staining fluorescence for visualizing microspore stages. These methods included staining with or without anther fixation, diluting DAPI in PBS with or without anther fixation, and using a combination of DAPI, EDTA, Triton X100, and PBS buffer, with or without a fixative. The study concluded that the most reliable approach involved diluting DAPI in PBS and fixing the anthers in Carnoy's fixative, providing an accurate visualisation of microspore stages.