水稻瘤矮病毒P12蛋白抗血清制备及其应用  

Preparation and application of antiserum against P12 protein of Rice gall dwarf virus

作　　者：孟媛[1,3] 辛星[2,3] 羊健[3] 张恒木[3] 王建明[2] 陈剑平[3] 

机构地区：[1]浙江师范大学化学与生命科学学院,金华321004 [2]山西农业大学农学院植物病理系,太谷030801 [3]浙江省农业科学院病毒学与生物技术研究所,浙江省植物有害生物防控国家重点实验室培育基地,农业部植物保护与生物技术重点开放实验室,浙江省植物病毒重点开放实验室,杭州310021

出　　处：《植物保护学报》2013年第5期413-417,共5页Journal of Plant Protection

基　　金：国家科技支撑计划(2012BAD19B03);公益性行业(农业)科研专项(201003031);浙江省自然科学基金(Y3090657)

摘　　要：为了进一步研究水稻瘤矮病毒(Rice gall dwarf virus,RGDV)P12的功能,将RGDV-GX P12编码区亚克隆至原核表达载体,并导入大肠杆菌诱导表达;重组的P12融合蛋白经Ni-NTA His·Bind Resins纯化后用于免疫小鼠制备抗血清,并进行Western blotting分析。结果显示,约41 kD大小的RGDV P12融合蛋白可在大肠杆菌中高效表达;利用该重组蛋白所制备的抗血清可特异地与融合表达和非融合的P12蛋白发生强烈的免疫学反应。利用该特异性抗血清在病毒粒子样品中检测不到P12蛋白,而在病株总蛋白样品中可检测到1条与预期大小一致的明显条带,表明RGDV P12是一种非结构蛋白。To further study the function of Rice gall dwarf virus （RGDV） P12, the encoding region of RGDV P12 was subcloned into the prokaryotic expression plasmid vector and transformed into Escherichia coli for inducible expression. The recombinant P12 fusion protein was purified with Ni-NTA His ~ Bind Resins and then used for preparing antiserum with immunized mouse. The resultant antiserum was used for Western blotting assay. The results showed that the RGDV P12 fusion protein with the size of approxi- mately 41 kD could be efficiently expressed in E. coll. The prepared antiserum could react specifically and strongly with both fusion and native P12 proteins. RGDV P12 was detectable by Western blotting with the specific antiserum in the expected size from the samples of infected rice plants but not from samples of virus particles, indicating that RGDV P12 was a viral nonstructural protein.

关 键 词：水稻瘤矮病毒 P12 原核表达 抗血清 

分 类 号：S435.111[农业科学—农业昆虫与害虫防治]