Expression of the gene for ribosome-inactivating protein, SoRIP2, as a tool for the evaluation of somatic embryogenesis in spinach

Abstract

High variability in regeneration capacity has previously been observed within a population of seedlings in several spinach cultivars. The cultivar “Matador” exhibits particularly low regeneration potential, and the majority of lines obtained in our previous study displayed a stable embryogenic capacity only for a limited period of time. In order to shorten the time required for embryogenic capacity assessment for individual lines, a model system for the rapid evaluation of embryogenic capacity was developed. This model system was based on the expression of a gene encoding spinach ribosome-inactivating protein (SoRIP2), which showed low expression levels in roots grown under non-inductive conditions. Induction of globular somatic embryos (SEs) resulted in a 285-fold increase in SoRIP2 expression that dropped to the control level beyond cotyledonary-stage SEs. The model system was tested by comparing the expression of SoRIP2 and the index of embryo-forming capacity (EFC), which integrates the frequency of regeneration and the mean SE number per root explant. Comparisons were always made within the same line, and the expression of SoRIP2 and the EFC index were determined 4 and 12 weeks after starting induction treatment, respectively. High positive correlations between SoRIP2 expression and EFC were obtained for the two factors that influenced embryogenic capacity the most: genotype (r2 = 0.81) and photoperiod (r2 = 0.92). The results indicate that the expression of SoRIP2 can be successfully used for early evaluation of regeneration capacity of individual lines, before SEs can be seen with the aid of a stereomicroscope, even 8 weeks earlier than by the conventional method.