辣椒疫霉菌致病毒素的纯化及其致病性的研究

Purification and Pathogenicity Detection of The Toxin Produced by Phytophthora capsici

通过硫酸铵沉淀法提取到疫霉菌的原始毒素。采用甜椒叶片注射法检测毒素的活性。采用两种方法对原始毒素进行纯化,用连续Bio-gel P-100柱层析法纯化能将毒素提至层析纯。在离子交换-分子筛层析法中,原始毒素依次经DEAE52-cellulose →CM32-cellulose→Sephadex G-75柱层析纯化后,经电泳检测为单一条带,说明达到电泳纯。采用SDS-PAGE法测定毒素的分子量为41.9 ku。对毒素的组分进行了测定,结果表明毒素中蛋白质含量为70.4%,糖含量为29.6%。毒素经蛋白酶K和胰蛋白酶处理后活性均下降,而用β-葡萄糖苷酶处理后活性变化不大,说明蛋白质亚基在P.capsici毒素活性中起着主要作用。

In this paper, original toxin was obtained from culture medium filtrate after (NH4)2SO4 precipitation. Toxin activity was evaluated by the degree of lesions on capsicum leaves after inoculation. Two methods were used to purify original toxin. The toxin could be purified to chromatography pure by a series of Bio-gel P-100 column separation. Using ion exchange and molecular sifting method, through DEAE52-cellulose, CM32-cellulose and SephadexG-75 column separation, original toxin could be purified to electrophoresis pure. Purified toxin showed itself only one bind on the picture of SDS-PAGE, and the molecular was 41.9 ku. The composition of the P. capsici toxin also be determined. The ratio of protein and sugar was 70.4% and 29.6% respectively. Toxin activity declined after treated by Proteinase K and Trypsin, but little changes after treated by β-glucosidase, which indicated that the protein plays an important role on the activity of P. capsici toxin.