Changes in surface structure during direct somatic embryogenesis in rice [Oryza sativa] scutellum observed by scanning electron microscopy

Direct somatic embryogenesis in rice scutella was studied by scanning electron microscopy paying attention to the morphological change of surface structure. Somatic embryos were formed when the scutella of immature embryos of cv. Nipponbare were cultured on MS media containing 2 mg L −1 2,4-D. Embryos were then transferred to the media containing 1 mg L −1 2,4-D and germinated on hormone-free media. The developmental stages of typical monocotyledonous somatic embryo were classified into proembryo, globular, scutellar and coleoptilar stages by referring to the terminology used for zygotic embryos. Each stage had a characteristic surface structure. The surface of proembryo was smooth. Fibrillar material was observed during the transition from proembryo to globular stage. Fibrillar material was then gradually replaced by a mesh-like structure at the globular stage. With subsequent development of globular embryo, the apical scutellum region became flattened and the ridge of mesh-like structure became lower. Development of scutellum followed by the emergence of coleoptile from the scutellar notch was observed at the scutellar stage. The coleoptile surface was covered with elongating cells. The coleoptilar stage was characterized by the growth of coleoptile and the development of root. However, aberrant somatic embryos which had leafy scutellum and multiple shoot meristem, were also observed.