| Hybrid sterility is one of the major barriers in utilization of heterosis in hybrid rice breeding. Therefore, it is important to clone corresponding genes for hybrid male sterility and to elucidate the molecular mechanisms underlying these important agronomic traits. The hybrid sterility gene Sa has been cloned, which consists of two adjacent genes, Sa M and Sa F and produces three of the factors, Sa F+,Sa M+, and Sa M-. Based on the genetic relationship, a two-gene/three-component interaction model has been proposed for the Sa-mediated hybrid male sterility. And an intercellular trafficking was hypothezed to explain the molecular mechanism of the model. In this study, I tried to test the possibility of intercellular trafficking of the three factors in microspore with immunohistochemical analysis to further validation the hypothesis. The main results of this study are as follows:(1)Three genetic transformation vectors with fusion tag were constructed and transgened into japonica rice T65(Sa M-/Sa F-) or indica rice E4(Sa M+/Sa F+),respectively.(2)The single site insertion of positive transgenic plants were obtained and confirmed by PCR and southern blotting.(3)According to pollen fertility and panicle phenotypes of transgenic plants,the T65 Sa M+-FLAG single copy transgenic plants were full-fertile; the T65 Sa M+-FLAG/Sa F+-HA single copy transgenic plants were full-sterile; and the E4 Sa M--FLAG single copy transgenic plants were semi-sterile.(4) However, the results of immunohistochemical analysis did not support intercellular trafficking hypothesis within the four small spore tetrad.Taken together, this study confirmed the two-gene/three-component interaction model in genetic context, but did not support the intercellular trafficking hypothesis for this model, which provides some clue to modify the Sa working model. |
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