棉 花 学 报 Cotton Science 2005，17(1)：29-32
Preliminary Probe to Binding Sites of a Verticillium dahliae Phytotoxin Abstract: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-1IgG), 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.
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