Evaluation of somatic embryo production during embryogenic tissue proliferation stage using morphology, maternal genotype, proliferation rate and tissue age of Pinus thunbergii Parl

To determine the optimal embryogenic capacity (somatic embryo production) of the selected elite nematode-resistant genotypes of Pinus thunbergii, variables such as embryogenic tissue (ET) morphology, maternal genotype, proliferation rate and tissue age were analyzed. ET morphology and histological evaluation of the proliferation stage showed a decrease in filamentous clump and protuberant surfaces and a decline in the acetocarmine-staining area, which indicates a decrease in somatic embryo production (SEP). Variations in cell physiology during the proliferation stage showed that SEP was positively correlated with soluble sugars and proteins, but negatively correlated with starch, peroxidase, and superoxidase. In addition, SEP was significantly (p < 0.001) affected by maternal genotype, tissue age and proliferation rate. Moreover, SEP was positively correlated with proliferation rate (r = 0.98, p < 0.001), but negatively correlated with tissue age (r =  − 0.95, p < 0.001). In general, the results suggest that SEP could be assessed in ET proliferation stages by the apparent cell morphology, histology, proliferation rate and tissue age, which provides novel insights for evaluating the ET maturation capacity (number of somatic embryos) during the proliferation stage of P. thunbergii somatic embryogenesis.