3.1 Expression of starch synthesis-related genes
In our previous study, the expressions of starch synthesis-related genes such as OsGPT2 , OsAGPL1 , OsAGPS1 , OsBEI andOsPho1 were highly correlated with the expression of CRCTin the leaf sheath of CRCT overexpression lines and knockdown lines (Morita et al., 2015; Morita et al., 2019). There are considerable differences in the starch content in the leaf sheath among rice varieties (Goda et al., 2016). Thus, in this study, the relationships between the expression levels of CRCT and the starch synthesis-related genes were analyzed in the six rice cultivars, Nipponbare, Koshihikari, Habataki, Takanari, Momiroman and Leafstar (Figure 1). Among these cultivars, Nipponbare was used for the production of transgenic lines. Koshihikari is the most popular cultivar in Japan. Habataki, Takanari and Momiroman are representative high-yielding varieties. However, the pattern of changes in the nonstructural carbohydrate (mainly starch) content in the leaf sheath and culm during the reproductive stage were different between Takanari and Momiroman (Yoshinaga et al., 2013). For example, the content of nonstructural carbohydrate in the leaf sheath and culm were significantly lower in Takanari than in Momiroman at the grain-filling stage. Leafstar is characterized as lodging resistant and has a high starch content in straw (Ookawa et al., 2010). Using the leaf sheath of these cultivars, the expression levels of CRCT were compared with the expression levels of starch synthesis-related genes by qRT-PCR. The expression levels of OsAGPL1 , OsAGPS1 and OsPho1highly correlated with that of CRCT. In contrast, weak or no correlation with CRCT expression were observed in OsGPT2 and OsBEI . However, when limited to Nipponbare, Koshihikari and Takanari, theOsBEI expression was correlated with the CRCT expression. While not apply to all, the expression level of CRCT can be a major determinant of natural variation in the expression level of multiple starch synthesis-related genes.
The relationship between the expression level of CRCT and starch content was also analyzed using these cultivars. In our previous study, it was clearly shown that starch content was greatly increased in theCRCT overexpression lines and decreased in the CRCTknockdown lines (Morita et al., 2015). However, a significant correlation was not observed between the starch content and the expression levels of CRCT in the leaf sheath among these rice cultivars (Figure 1). This tendency was the same even when limited to the cultivars showing the correlation between the OsBEIexpression and the CRCT expression.