Premise of the study: Protocorm-like bodies (PLBs) of orchids are important in orchid micropropagation and outwardly resemble somatic embryos in form and development. To determine whether PLBs are truly embryogenetic, we compared PLBs with somatic embryos and zygotic embryos to determine whether they had similar surface molecules and whether hydroxyproline-rich glycoprotein (HRGP) inhibitors similarly alter their growth.
Methods: Embryogenic calluses (ECs), zygotic embryos, and protocorms were collected for histological and histochemical studies with light microscopy. The presence of JIM11 and JIM20 epitopes was determined using immunodot blots and immunolocalization procedures. The importance of wall proteins in the formation of PLBs was investigated using 3,4-dehydro-l-proline (3,4-DHP), an inhibitor of HRGP biosynthesis.
Key results: At the early stages of PLB formation, the cytoplasm of the globular cell clusters and meristemoids took on a vacuolated appearance. Starch granules and protein bodies appeared, albeit transitory in nature. Positive localizations of JIM11 and JIM20 were noted in the embryogenic culture and developing PLBs similar to zygotic embryos. The inclusion of an inhibitor to HRGPs inhibited PLB formation.
Conclusions: This study demonstrates that during the early stages of PLB formation, the cells show cytological characteristics and cell wall markers similar to zygotic embryo development, justifying the statement that PLBs are indeed somatic embryos of orchids. Thus, these results suggest that PLBs could be used as an experimental embryological system for physiological or molecular characterization.
Keywords: JIM11; JIM20; Orchidaceae; Phalaenopsis; hydroxyproline-rich glycoproteins; meristemoids; protocorm-like bodies; somatic embryo.