Changes in biochemistry and histochemical characteristics during somatic embryogenesis inOrmosia henryiPrain

ABSTRACT Mature embryos were used as an explant for embryogenic callus (EC) induction, and then EC was further developed to form somatic embryos during somatic embryogenesis (SE) of Ormosia henryi Prain; however, some mature embryos could induced non-embryogenic callus (NEC), browning callus (BC) or snowflake callus (SC). These phenomena might be related to the biochemical and histochemical differences during somatic embryo induction. The present study was conducted to analyze the biochemical events and histochemical changes at different SE stages in 0. henryi . The contents of soluble sugar, starch, soluble protein, H 2 O 2 , and endogenous hormones and the activities of polyphenoloxidase (PPO), superoxide dismutase (SOD), ascorbate peroxidase (APX), peroxidase (POD) and catalase (CAT) were measured at different SE stages, such as EC, globular embryo (GE), and cotyledon embryo (CE), and in abnormal tissue, such as NEC, BC, and SC. The results showed that the contents of soluble sugar and starch; the activities of PPO, SOD, APX and POD; and the ratios of indole-3-acetic acid/abscisic acid (IAA/ABA), IAA/gibberellins (IAA/GAs), auxin /GAs (AUX/GAs), and AUX/ABA decreased gradually at different SE stages. In contrast, the contents of soluble protein, H 2 O 2 , all endogenous hormones gradually increased. However, CAT activity and the ratios of IAA/cytokinins (IAA/CKs), AUX/CKs, ABA/CKs, and GAs/CKs first increased and then decreased. The high contents of GAs and ABA, high ratios of ABA/CKs and GAs/CKs and low ratios of IAA/ABA, IAA/GAs, AUX/GAs and AUX/ABA were responsible for the inability of the callus to form EC. The low enzyme activities, low contents of energy substances and H 2 O 2 were related to NEC formation. The high contents of soluble sugar, H 2 O 2 , AUX, CKs and PPO activity and the low content of soluble protein were the basic causes of BC formation. The high-energy substances contents and low activities of SOD and POD facilitated SC formation. Histochemical observation showed that starch granule staining gradually lightened with SE development, but protein granules were darkly stained. Compared with EC, starch and protein granules were stained darker in SC, and lighter in NEC and BC. These results showed that energy substances were the material basis of SE, which affected enzyme activities, regulated reactive oxygen species (ROS) metabolism, and thus regulated the morphogenesis and development of somatic embryos. In addition, the contents and ratios of endogenous hormones affected the dedifferentiation, dedifferentiation and embryogenesis of somatic cells. To induce EC from mature embryos and further develop their formation into somatic embryos, it is necessary to adjust the energy supply and hormone ratio in the medium. One-sentence summary Somatic embryogenesis and abnormal callus tissues formation of Ormosia henryi Prain were associated with energy substances, reactive oxygen species, enzyme activities and endogenous hormones, as well as histochemical characteristics.