水稻高效再生体系的建立及其对大豆Em基因的转化  

Establishment of Efficient Rice Regeneration System and Transformation of Soybean Em Gene

作　　者：刘国宝[1] 刘昀[1] 郑易之[1] 

机构地区：[1]深圳大学生命科学学院,深圳市微生物基因工程重点实验室,广东深圳518060

出　　处：《西北植物学报》2008年第4期686-691,共6页Acta Botanica Boreali-Occidentalia Sinica

基　　金：深圳市科技计划项目(200440);国家科技部转基因专项研究(ZJY03-A-18-03)

摘　　要：以粳稻丰达1号成熟胚为外植体,建立了适合水稻转化的高效再生体系,通过农杆菌介导法将大豆Em基因转化水稻.结果表明:以NMB+500 mg·L-1脯氨酸+250 mg·L-1谷氨酰氨+300 mg·L-1水解酪蛋白+2 mg·L-12,4-D为作诱导培养基诱导愈伤组织,其诱导率为97%;愈伤组织分化率为65%;农杆菌介导法转化水稻愈伤组织,获得了26株再生植株;随机选取其中4株候选转基因水稻幼苗进行PCR筛选,初步证明大豆Em基因已整合到水稻基因组DNA中;RT-PCR检测结果表明,转入的外源大豆Em基因在转基因水稻中得以表达.本实验成功地建立了适合水稻转化的高效再生体系,为通过基因工程技术培育水稻新品种奠定了基础.By using mature embryo of Japonica rice ‘Fengda 1＇ as an explant,a highly efficient regeneration system suitable for rice genetic transformation was established, By transferring the soybean Era（group 1 LEA gene）gene into rice through Agrobacterium-mediated method. We approved the reliability of the established regeneration system. The results showed that YI（NMB＋500 mg · L^-1 Pro＋250 mg · L^-1 Gin＋ 300 mg · L^-1 hydrolyzed casein＋2 mg · L^-1 2,4-D）was used to induce callus. The inducement rate reached 97 % and the differentiation rate was 65 %. The expression vector of the soybean Em gene was constructed. Screening with the hygromycin after the vector has been delivered into the rice callus through Agrobacteri- urn-mediated method and 26 regeneration plants had been obtained; 4 transgenic rice seedling candidates were chosen randomly and PCR （polymerase chain reaction） was carried out with the specific primer to amplify anti-hygromycin gene and soybean Em gene respectively. It preliminarily proved that the soybean Em gene had already been integrated into the rice DNA genome. Furthermore, the result of RT-PCR demonstrated the expression of exogenous soybean Em gene in the transgenic rice. These results suggested that an efficient regeneration system suitable for rice transformation had been set up, which provides the foundation for breeding new rice species by genetic engineering technology.

关 键 词：大豆Em基因 转化体系 水稻 农杆菌介导 LEA蛋白 

分 类 号：Q789[生物学—分子生物学]