Alain Deloire

Arrest of somatic embryo development in grapevine: histological characterization and the effect of ABA, BAP and zeatin in stimulating plantlet development

In an effort to understand the causes of arrest of somatic embryo development, generally observed in grapevine (Vitis sp.), histological studies were undertaken, using two cultivars (CH76 and 41B) which differ in their ability to develop into plants. Embryos with a high conversion rate (70%; CH76) formed a well-structured and functional shoot apex between two thread-like cotyledons. In contrast, embryos with a low conversion rate (10%; 41B) formed a normal root apex but lacked a well-structured shoot apex and developed a wide range of aberrant forms in the intercotyledonary area: uncontrolled cellular proliferation, formation of adventitious buds, over-growth of cotyledonary or leaf meristems. ABA increased the conversion rate of 41B embryos from 10% to 20%, but failed to improve embryo morphology. Zeatin and BAP promoted growth of 41B somatic embryos, but generated a high level of abnormalities and failed to improve conversion rate. Applied in combination with ABA, these PGRs increased the frequency of cotyledonary embryos, but decreased the conversion rate.

Somatic embryogenesis from grapevine cells. I-Improvement of embryo development by changes in culture conditions

In conventional culture conditions without auxin, somatic embryos arising from suspension cultures of grapevine rootstock 41B (Vitis vinifera cv. Chasselas x Vitis berlandieri) are arrested at the heart stage of development. Starting from indications that inhibitors excreted in the culture medium could be responsible for this arrest, new culture conditions based on daily subculturing embryos in fresh medium have been successfully used to obtain full embryo development. From this technique, a microassay was devised for screening small amounts of extracellular molecules as potential inhibitors of embryonic development. Our results show that extracellular macromolecules of molecular weight higher than 10 kDa are likely involved in the inhibition of caulinary meristem initiation. However, other factors obviously cooperate to inhibit embryo development in conventional culture conditions

Extracellular protein patterns of grapevine cell suspensions in embryogenic and non-embryogenic situations

Abstract Somatic embryos produced from suspension cultures of grapevine rootstock 41B (Vitis vinifera cv. Chasselas × Vitis berlandieri) are blocked at the heart stage of development. Previous results have shown that extracellular macromolecules of molecular weight > 10 kDa, are likely involved in this inhibition (P. Coutos-Thevenot et al., Plant Cell Tiss. Org. Cult., 23 (1992) 125–133). The extracellular proteins and glycoproteins secreted by embryos differentiating in the absence of auxin have been compared to those secreted by undifferentiated cells growing in the presence of auxin. Specific proteins and glycoproteins (66, 62, 56, 51, 48, 36, 32, 25 and 10 kDa), under negative control of auxin, are secreted during embryogenesis. Thus, somatic embryogenesis in grapevine presents several similarities with the carrot model system as far as the influence of auxin on extracellular protein patterns and embryo differentiation is considered. This conclusion has been reinforced by showing that, as described in the carrot system, 36-kDa glycoprotein excreted by grapevine embryos is a cationic peroxidase whereas the 10-kDa protein immunoreacts with an antibody directed against a maize lipid transfer protein. Study of extracellular protein patterns of a Chardonnay CH76 cell strain, strongly affected in its embryogenic response, showed significative differences in the excretion of some specific proteins compared to the 41B cell line.