棉花黄萎病菌毒素结合位点初探  

Preliminary Probe to Binding Sites of a Verticillium dahliae Phytotoxin

作　　者：刘凤权[1] 胡白石[1] 

机构地区：[1]农业部病虫监测与治理重点实验室南京农业大学植保学院,南京210095

出　　处：《棉花学报》2005年第1期29-32,共4页Cotton Science

基　　金：国家自然科学资金(39800090)

摘　　要：以纯化的棉花黄萎病菌毒素(PLPC)免疫新西兰白兔制备了PLPC特异性抗血清。将PLPC(15μg·ml 1)用不同浓度的抗体(1～20μg·ml 1IgG)吸附后处理泗棉3号的切根苗,毒素所引起的症状都有不同程度的减轻。表明所制备的抗体在与毒素发生特异性免疫学反应的同时,可部分封闭毒素分子上与毒素受体结合的位点。利用竞争ELISA测定了泗棉3号幼苗子叶的质膜制剂与PLPC的结合活性。结果表明,质膜制剂与毒素结合后能部分阻断毒素与其抗体的免疫学反应,即质膜制剂中含有毒素的结合位点。分别用胰蛋白酶和煮沸处理质膜制剂后,质膜制剂对毒素与其抗体的反应的抑制作用消失,初步表明质膜制剂中与毒素结合的是蛋白质。A phytotoxic protein-lipopolysaccharide complex (PLPC) produced by Verticillium dahliae of cotton was purified by Fast Protein, Peptide and Polynucleotide Liquid Chromatography (FPLC). A specific polyclonal antibody against the phytotoxin was prepared by injecting the New Zealand rabbits with the purified PLPC. When the PLPC (15 μg·ml^(-1)) was absorbed by the prepared antibody of different concentration (1～20 μg·ml^(-1) IgG), the wilting symptom of cotton seedlings (cv. Simian 3 ) caused by the PLPC was alleviated to different degree, and the disease index was decreased from (13.4%) to 56.2%. The result showed that the prepared antibody not only immunologically react with the PLPC, but also partly block some sites (groups) of PLPC molecular which can recognize the binding sites on the cotton plant cells. The binding of the plasma membrane preparation of cotton cotyledon with the PLPC was determined by the indirect competitive ELISA. Results demonstrated that the immunological reaction of the PLPC and the antibody can be inhibited by the plasma membrane preparation (the inhibition varied from 8.2% to 54.8%), which hints that the plasma membrane preparation contains the binding sites of the PLPC, and the binding sites of PLPC maybe exist in the plasma membrane preparation of seedling cotyledons of cotton. When the plasma membrane preparation was treated by 5 μg·ml^(-1) L-trypsin for 30 min or boiled at 100℃ for 15 min, the immunological reaction of the PLPC and the antibody can not be blocked perfectly by the membrane preparation, which shows that some proteins in plasma membrane preparation are probably responsible for binding with the PLPC.

关 键 词：质膜 棉花黄萎病 毒素 切根苗 子叶 幼苗 新西兰白兔 结合位点 结合活性 纯化 

分 类 号：S435.621[农业科学—农业昆虫与害虫防治]