A Study On Transformation Of The HLF Gene And SB401 Gene Into Rice Mediated By Agrobacterium

Rice (Oryza sativa L.) is one of the most important food crops in the world,and it is also staple food of more than fifty percent population in China.As staple food of majority, high and stable yield of rice is first paid moreattention by people. With the living standard being improved, more and morepeople focus on the nutritional value of rice especially the content of proteinand lysine.We can improve multiple traits of rice such as protein quality at the sametime by conventional breeding approaches or sexual crossing betweentransgenic plants, but they both cause long breeding cycle and low efficient.Introducing multiple genes which present in the same T-DNA region of oneplasmid into target rice is an efficient approach to improve multiple traitsof rice simultaneously. In this study, hLF gene and a lysine-rich protein(sb401) gene which are controlled by a rice endosperm-specific expression(PGlu) promoter were transferred into two rice varieties viaAgrobacterium-mediated genetic transformation. The two genes were carried inthe same T-DNA region of plasmid p1304. The results were as follows:1. By comparison of the condition of the explants inducement and the stateof the callus, a conclusion was drawn that the inducement rate of the immatureembryos of Zhonghua 15 and Jingkang No. 5 is the highest. And the state of callusderived from their immature embryo was also the best. The inducement rate ofZhonghua 15 was higher than that of Jingkang No. 5, while callus derived fromJingkang No. 5 was more appropriate for suspension culture than that derivedfrom Zhonghua 15 when it comes to the state of the callus.2. The suspension cell lines of Jingkang No. 5 can be established within50 days. Through the measurement of the growth characteristics and theobservation of the differentiation performance of japonica rice Jingkang No. 5at different phases, we identified the best interval for every subculture was7-10d, and the prime time for the regeneration of the suspension cells was the middle phase.3. The selection and regeneration results of Zhonghua 15 callustransformants and Jingkang No. 5 suspension cells indicates that the bestselection concentration of hygromycin for Zhonghua and Jingkang No. 5 is 50mg/Land 30mg/L, respectively.4. After callus of Zhonghua 15 was transformed and selected withcephalosporins and hygromycin, regenerated seedlings were obtained and grewmature. The PCR assay initially proved that transgenic plants containingforeign genes of hLF and sb401 was obtained.