| Rice(Oryza sativa) is the world’s major staple food, study of rice male sterile relate genes beneficial to understanding the molecular mechanism of male reproductive development. Previous studies in our laboratory found that rice OsRacD Rop protein associated with the fertility of rice. To study its mechanisms, through the yeast two-hybrid screening panicle cDNA library,several protein-coding genes was isolated from which have interaction with OsRacD, including OsRhoGDI2 in this study. For the identification of OsRhoGDI2 genes function, especially the relationship with rice male development, the laboratory was constructed gene overexpression vector in transgenic rice, and screening of each generation. Based on the previous study, overexpression of OsRhoGDI2 gene in transgenic rice to measure T3 and analysis, aims to identify the function of the gene and lays the foundation of clarifies the connection of this gene and Os RacD in development of rice.OsRhoGDI2 genetically modified T3 rice was screened by PCR technique, analyzed the level of expression of OsRho GDI2 gene in transgenic rice plants with qRT-PCR technology, showed that 90% above the T3 generation of rice was positive plants, and levels of gene expression in transgenic rice plants were higher than those in the control group, increases of 5 to 26 times. Indicates that OsRhoGDI2 expression in transgenic rice plants in the T3 generation has become more stable, can be used as a functional dissection OsRhoGDI2 material.Selection of panicle/flower from which in stamen formation as well as 8 typical stages of rice microspore development in wild-type and transgenic plants respectively, making longitudinal sections and transverse section of paraffin, and was observed. Results showed genetically modified rice have more spikelet each panicle than that in wild-type plant, and its distribution is more intensive. Microspore development process of transgenic rice and wild-type rice pre-meiosis, meiotic have not significant differences in cytology, but in the late microspore development, transgenic rice microspore atrophy, resulting in part of maturate pollen collapsed abnormal. These differences suggest that overexpression of OsRhoGDI2 gene significantly affects the differentiation of rice spikelets, later development of rice microspore and lead to the anther dysplasia, pollen morphological abnormalities.To test for mature pollen viability from transgenic rice and wild rice, using pollen staining and pollen germination in vitro. Results showed that compared to wild-type rice, rice pollen fertility rates decreased significantly, fell to 79% from 95% in control and genetically modified rice and the pollen germination rates in vitro is low also. Show that overexpression of OsRhoGDI2 gene in rice pollen activity decreased. Statistical analysis of mature rice agronomic, similar to the discovery which detected from T2 transgenic rice, the T3 generation of transgenic rice has longer panicles, increase grain number per panicle but seed set is significantly lower than wild-type rice. Paraffin sectioned slides together with viability pollen test results suggest that overexpression of OsRhoGDI2 affect the late microspore development, lead to dysplasia anthers, pollen viability decreased and low seed setting rate in transgenic rice.This research based on laboratory previous studies, screened the generation of OsRhoGDI2 transgenic rice, analyzed the overexpression level of OsRho GDI2 gene in transgenic rice, compared the differences between transgenic rice and wild type rice spike differentiation and development of small spores and their agronomic traits for statistics analysis, not only confirmed that the gene associated with rice panicle length and grain number, and found that the gene functions involved in important processes of panicles development and post-development of small spores. For more in-depth follow-up study in OsRho GDI2 and the contact with OsRacD in growth and fertility of rice provide clues and evidence. |
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