ISHS Acta Horticulturae 472: XVII International Symposium Virus and Virus-Like Diseases of Temperate Fruit Crops
DETECTION OF APPLE CHLOROTIC LEAF SPOT VIRUS AND APPLE STEM GROOVING VIRUS BY DOT-IMMUNOBINDING ASSAY |
| Authors: | Q. Wang, Y. Shi, J. Yang, Q. Sun |
| Abstract:
Three thousand samples were collected from 72 cultivars, 10 stocks and 6 indicator plants infected with viruses and then apple chlorotic leafspot and apple stem grooving viruses were detected by dot-immunobinding assay (DIBA). Plant tissue was extracted with phosphate buffer saline (PBS), 0.05% Tween-20, 2% polyvinylpyrrolidone (PVP) and 0.2% bovine serum albumin (BSA) (w/v=1:2 for tree leaves, and w/v=1:1 for test tube seedlings). Equal amount of chloroform was added for centrifugation. Two microliters of crude extract and 25 μl of horseradish peroxidase scale protein A(HRP-SPA) and 25 μl of antibody (1/4000 to 1/6000 dilution) were used in DIBA. DIBA is simple, rapid (5 h), and consistent for a whole growing period. It is economical and convenient. The amount of antigen, antibody and HRP-SPA required for DIBA is only 1%, 2–3% and 12.5% of that for ELISA respectively. DIBA can be directly read without any instruments. Nitrocellulose membrane used for DIBA can be stored for later reading. | |
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