番茄SlWDR141基因的克隆及功能研究

摘要

为研究番茄WD40蛋白基因(SlWDR141)的功能,对SlWDR141蛋白进行生物信息学分析,发现SlWDR141蛋白在进化过程中高度保守,与土豆的同源性最高.在克隆SlWDR141基因全长序列的基础上,构建了亚细胞定位和酵母双杂交表达载体,证实SlWDR141蛋白主要定位在细胞核中,与DDB1具有相互作用.实时荧光定量PCR(real-time PCR)分析SlWDR141基因在组织中的表达谱,发现它在各个组织中均有表达,但在根中的表达量最高.为研究该基因的生物学功能,进一步构建了SlWDR141基因的干涉载体,用农杆菌介导法转化番茄得到转基因阳性番茄,荧光定量PCR鉴定出3个表达量下调的独立转基因株系.NaCl胁迫和Pst DC3000侵染野生型和转基因植株发现,转基因植株相对于野生型植株抗盐性和抗病性均下降,表明SlWDR141基因对盐胁迫和细菌侵染均起正调控作用.%To elucidate the functions of tomato WD40 gene SlWDR141, bioinformatics analysis for SlWDR141 protein was carried out.We found that SlWDR141 protein is highly conservative in evolution and has the highest homology with potato.By cloning the full-length sequence of SlWDR141 gene, the expression vectors for subcellular localization assay and yeast two-hybrid analysis were constructed.We confirmed that SlWDR141 protein is mainly localized in nucleus and SlWDR141 physically interacted with tomato DDB1.Real-time PCR analysis indicated that SlWDR141 gene expressed in all tested tissues but in root is the highest.In order to study the biological function of the gene, the interference vector of SlWDR141 gene was further constructed.Transgenic tomato was obtained by Agrobacterium-mediated transformation, and three independent transgenic lines were identified by fluorescence quantitative PCR Salt stress and Pst DC3000 infection analyses demonstrated that transgenic plants are less tolerant to the salt stress and pathogenic bacteria than wild-type plants, indicating that SlWDR141 gene plays a positive role in response to biotic and abiotic stresses.