Anther culture and cold treatment of floral buds increased symmetrical and extra nuclei frequencies in soybean pollen grains

Androgenic response is characterized by a multinucleate or multicellular stage of pollen development. Histological sections stained with toluidine blue and squashes in propionic-carmine and in 4-6-diamidino-2-phenylindole (DAPI) were used for serial observations (0, 14 and 28 days) in soybean pollen grains from cultured anthers and floral buds submitted to cold treatment at 4 °C. In a total of 62,536 pollen grains, it were observed general averages of 2.06% of pollen grains with two symmetrical nuclei and of 1.41% pollen grains with typical extra nuclei (i.e. additional nuclei with typical morphology). Symmetrical and extra nuclei frequencies increased in both treatments but only the number of pollen grains with typical extra nuclei increased significantly with time of exposure to treatments. In addition, 8.59% of multinucleate pollen grains were recorded with atypical nuclei, smaller than vegetative or generative-types and with a fragmented shape. The frequency of these grains increased significantly with time of exposure to treatments. Thus, soybean multinucleate grains occurrence was not an exclusive response to culture. These preliminary results point to the need of further studies to clarify the relationship between typical and fragmented extra nuclei with both androgenesis and programmed cell death.