Abstract:

The aim was to study the technology which detected Odontoglossum ringspot virus (ORSV), which was one of the most important and worldwide viruses attacking orchids. Chinese Cymbidium plants which showed disease symptoms were collected from Zhangzhou, Fujian. One pair of specific primers was designed for amplification of the coat protein (CP) gene from the samples infected with ORSV. The open reading frame encoding CP of ORSV isolate obtained from Zhangzhou, Fujian is 477 bp, encoding a 18.0 kD protein with 158 aa. The expected CP gene was then inserted into the pET-29a (+) vector for the prokaryotic expression. And the aimed protein was purified and used to immune the rabbit for antiserum preparation. According to the result of ID-ELISA analysis, specific rabbit anti-ORSV serum was prepared with a high titre of 1:25600, a working concentration of 1:12800, and a sap sensitivity of 0.195 mg/mL. Western blot analysis confirmed that the antiserum reacted strongly and specifically to the CP of ORSV. There were no obvious cross reactions between the antiserum and 11 kinds of homologous or heterologous viruses such as Tobacco Mosaic Virus (TMV).