论文部分内容阅读
采用ＤＥＡＥ－Ｔｏｙｏｐｅａｒｌ离子交换、Ｂｕｔｙｌ－Ｔｏｙｏｐｅａｒｌ疏水层析、ＣＭ－Ｔｏｙｏｐｅａｒｌ离子交换、ＳｅｐｈａｃｒｙｌＳ１００凝胶过滤和ＦＰＬＣＭｏｎｏＱ等步骤，从非亲和性稻瘟菌侵染稻叶中纯化了两种诱导性脂氧合酶，即ＣＭ－Ｌｏｘｌ和ＣＭ－Ｌｏｘ２。ＳＤＳ－ＰＡＧＥ检测结果表明：ＣＭ－Ｌｏｘ１和ＣＭ－Ｌｏｘ２为单链多肽，它们的分子量分别为９８ｋｄ和１０２ｋｄ。利用微量免疫法制备了ＣＭ－Ｌｏｘ１和ＣＭ－Ｌｏｘ２的抗体，制备的抗体效价达到１∶１０００倍，能检测０．５～２．０ｎｇ的抗原。免疫印迹杂交证明ＣＭ－Ｌｏｘ１和ＣＭ－Ｌｏｘ２之间存在密切的血清学关系。此外，Ａｎｔｉ－ＣＭ－Ｌｏｘ１和Ａｎｔｉ－ＣＭ－Ｌｏｘ２与水稻中另一稻瘟菌诱导性脂氧合酶ＲＬＬ以及发芽种子出现的ＲＬ－２也存在交叉反应。
Two kinds of induced lipoxygenases were isolated from rice inoculated with non-affinity Magnaporthe grisea using DEAE-Toyopearl ion exchange, Butyl-Toyopearl hydrophobic chromatography, CM-Toyopearl ion exchange, Sephacryl S100 gel filtration and FPLCMonoQ. Synthase, namely CM-Loxl and CM-Lox2. The results of SDS-PAGE showed that CM-Lox1 and CM-Lox2 were single-chain polypeptides, and their molecular weights were 98kd and 102kd, respectively. Antibodies CM-Lox1 and CM-Lox2 were prepared by microimmunization. The titer of the prepared antibody reached 1: 1000 times, which can detect 0.5 ~ 2.0ng antigen. Western blot hybridization demonstrated a close serological relationship between CM-Lox1 and CM-Lox2. In addition, Anti-CM-Lox1 and Anti-CM-Lox2 also cross-reacted with another Magnaporthe grisea-induced lipoxygenase RLL in rice and RL-2 in germinating seeds.