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细菌脂多糖由O-抗原,核心多糖和脂质A三部分组成,脂质A是细菌内毒素活性的根源.植物脂多糖与细菌脂多糖组成基本相似,但不具有微生物脂多糖的高毒性.目前,对植物脂多糖的合成机制缺乏最基本的认识,因此研究植物脂多糖的合成具有重要的科学价值.作者在水稻基因组中发现一个含有大肠杆菌Ec Lpx A功能结构域的基因,命名为OsLpxA,该基因催化水稻脂质A合成的第一步反应.本研究用水稻(Oryza sativa subsp.Japonica)苗期的叶片为材料,提取总RNA并以此为模板,用反转录PCR扩增OsLpxA基因的si RNA靶序列,并将其连接到表达载体p TCK303上,构建了RNA干扰载体p TCK303-OsLpxA-RNAi.将该载体通过农杆菌介导法转化水稻,共获得59株具有潮霉素抗性的转化苗.然后利用潮霉素的特异性引物对全部抗性苗进行PCR检测,其中有38株转化苗呈阳性,表明潮霉素标记基因已整合到了水稻的基因组中.最后,利用定量PCR检测38株阳性苗中OsLpxA基因在转录水平表达的变化.结果表明,其中27株阳性苗中导入的OsLpxA基因RNA干扰结构成功地降低了目的基因的表达.该结果为后续对OsLpxA基因功能的研究提供参考.“,”Bacterial lipopolysaccharide consists of 3 parts:O-antigen, core oligosaccharide and lipid A.The lipid a domain is responsible for the toxicity of Gram-negative bacteria.The composition of plant lipopolysaccharides is similar to that of bacterial lipopolysaccharides, but plant lipopolysaccharides do not have the high toxicity as microbial lipopolysaccharides do.At present, there is no basic understanding of the formation mechanism of plant lipopolysaccharides.Therefore, it is of important scientific value to study the synthesis of plant lipopolysaccharides.In this study, a gene that contains an Ec Lpx A-like functional domain was found in rice genome.The gene was named as OsLpxA, which catalyzed the first step of lipid A biosynthesis.Oryza sativa (subsp.Japonica) was used as research materials in this study.The total RNA was extracted as a template from the leaves of rice seedlings, then the si RNA target fragment of OsLpxA gene was amplified by RT-PCR and ligated to the expression vector pTCK303 to construct the RNA interference vector p TCK303-OsLpxA-RNAi.The recombinant vector was transf ormed into rice by the method of agrobacterium mediated transformation and 59 transgenic rice plants with hygromycin resistance were obtained.Then, all resistant plantlets were detected by PCR using hygromycin specific primers, 38 transformants were positive, indicating that hygromycin marker gene had been integrated into the rice genome.Finally, fluorescent quantitative PCR was used to detect the expression of OsLpxA gene at the transcriptional level in 38 positive plantlets.The results showed that the Si RNA structure of exogenous gene OsLpxA in 27positive plantlets successfully reduced the expression of endogenous gene OsLpxA, which would lay the foundation for further research on the function of OsLpxA gene.