Construct And Transformation Of OsELF3 RNAi Vector In Oryza Sativa

Rice is one of the most important food crops to humans. Heading stage is one of the important agronomic characters for rice, which decided the ecological zones and regional adaptability of rice. One of the breeding objectives for rice breeders is to breed the rice varieties with precocious high-yield. Plant growth period regulation is a complicated process, involving many genes. A gene named ELF3 in arabidopsis is a positive regulative factor for circadian clock. The fuction of ELF3 is to regulate 24-hour biological rhythms and seasonal rhythms in order to maintain the viability of biological rhythms. The mutation of this gene lead to the loss of function for light cycle sensitivity and rhythm control. And the phenotype is performing flowers no matter in long or short daylight exposure conditions. Via blasting online, we found that the coding protein of OsELF3 in rice is highly similar to the coding protein of ELF3. So we speculate that the OsELF3 has the function of regulating rice heading stage. As a new biological technology, RNA interference technology (RNAi) can suppress specific gene expression by the regulation of gene transcription. This method has the advantage that only change the target gene without changing the genetic background of the plant itself. This make RNAi become a kind of simple and effective technology for gene knockout.This study has successfully constructed the expression vector(named pOsActl-OsELF3-RNAi) for RNAi to rice OsELF3 gene. And this recombined expression vector with OsELF3 is transmitted to Zhonghua 11 and Nipponbare by agrobacterium mediated transgenic technology. And several transgenic plants have been obtained. This may establish the foundation of further research for OsELF3 gene. The main results of this study are as follow:1 According to the OsELF3 gene cDNA sequence from Gramene, pairs of primer was designed to amplicate the 500bp RNAi-Sence and RNAi-Antisence fragments, respectively. 2 OsELF3 gene with RNAi fragment was cloned to expression vector pOsAct2-1-nos and the RNAi expression vector named pOsAct2-OsELF3-RNAi was constructed successfully.pOsAct2-OsELF3-RNAi is regulated by Actin2 promoter and the selection markers is Hpt gene.3 The RNAi expression vector (OsELF3-pOsAct2-RNAi) was transformed into the embryo callus of Zhonghua Hand Nipponbare by agrobacterium mediated transgenic technology. Through tissue culturing and resistance screening, the regenerate plants was obtained.4 The result of hygromycin testing for vitro blade and amplification of Hpt genes PCR by for To transgenic line indicate that the OsELF3-pOsAct2-RNAi was inserted into the rice genome.5 Compaired the positive RNAi T1 transgenic line with CK(Zhonghua Hand Nipponbare) shows that:the heading stage of T1 generation line are delyed. Statistical analysis indicate that this discrepancy is significantly different. The pOsAct2-OsELF3-RNAi transgenic line become an important material for advanced research for OsELF3 gene.