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为获得无任何抗生素抗性基因的抗病转基因水稻新品系,将克隆自甜椒的两亲性蛋白AP1基因与潮霉素抗性选择标记基因(HPT)分别构建位于同一农杆菌双元载体上的两个独立的T-DNA区中,并经农杆菌介导,将其导入江苏省推广的两个粳稻品种广陵香粳和武香粳9号中,获得了一批转基因水稻植株。PCR和Southern杂交分析表明,AP1基因和HPT基因已同时导入受体基因组中,并从共转化植株的自交后代中筛选到了无HPT基因的转AP1基因水稻植株。结合田间农艺性状考察,选育了多个无抗性选择标记基因的纯合转AP1基因水稻新品系;抗病性鉴定结果表明,转AP1基因水稻对稻白叶枯病的抗性与未转化对照相比有显著提高,部分转基因水稻对纹枯病的抗性与未转化对照相比也有一定程度的提高。
In order to obtain a new line of disease-resistant transgenic rice without any antibiotic resistance gene, the AP1 gene cloned from sweet pepper and the hygromycin resistance selection marker gene (HPT) were respectively constructed on the same Agrobacterium binary vector Mediated by Agrobacterium tumefaciens and introduced into two varieties of rice Guangling Xiangjing and Wuxiangjing 9 which were popularized in Jiangsu Province. A number of transgenic rice plants were obtained. PCR and Southern blot analysis showed that the AP1 and HPT genes were introduced into the genome of the recipient at the same time and the AP1 transgenic rice plants without HPT gene were screened from the selfed crosses of the co-transformed plants. Combining with the field agronomic traits, a number of homozygous transgenic rice lines of AP1 gene with multiple resistance-free selectable marker genes were selected. The results of disease resistance identification indicated that the resistance and non-transformation of AP1 transgenic rice to rice bacterial leaf blight Compared with the control, there was a significant increase, some of the resistance of rice to sheath blight of transgenic rice compared with the untransformed control also increased to some extent.