The purpose of this study was to determine if abscisic acid (ABA) was related to genotype differences of orchardgrass (Dactylis glomerata L.) for producing somatic embryos from leaf sections cultured in vitro. Leaf sections of one embryogenic (Embryogen-P) and one nonembryogenic (I- 39) genotype were exposed to various concentrations of ABA in the culture medium for differing lengths of time to determine the effect of exogenous application. Endogenous levels of the two innermost leaves were analyzed by high performance liquid chromatography (HPLC) and enzyme linked immunosorbent assay (ELISA) and compared in one embryogenic and two nonembryogenic genotypes. Comparisons were also made between basal (0-3 cm) and distal (3-6 cm) portions of the innermost leaves of one embryogenic and one nonembryogenic genotype.
In the embryogenic genotype, somatic embryogenesis was enhanced by a low level (1 μM) of ABA, while higher levels (10-100 μM) inhibited embryogenesis. Duration of exposure to ABA was also found to be significant, with an initial 3 day exposure period being most effective for enhancing embryo formation.
The nonembryogenic genotype showed no response to ABA application in any of the treatments.
Quantification studies indicated a significant difference in ABA concentrations between the embryogenic and nonembryogenic genotypes. When quantified by immunoassay, the embryogenic genotype contained more ABA in basal and distal sections of inner leaves than did the nonembryogenic genotype, 1-39; however, the second leaf of the nonembryogenic genotype contained more ABA than the embryogenic genotype. High performance liquid chromatography quantification results of combined first and second leaves indicated a significantly higher endogenous level of ABA in the nonembryogenic genotypes than in the embryogenic genotype. There was no significant difference between the two nonembryogenic genotypes.
Abscisic acid immunoassay results indicate that ABA levels decreased as much as 93% in both embryogenic and nonembryogenic genotypes during a 2 week culture period after initial plating.
These results suggest that both concentration and duration of application of ABA influence somatic embryo formation
