蝴蝶兰2种病毒的同步检测及其CP融合反义表达载体构建.

【Objective】Cymbidium mosaic virus(CyMV) and Odntoglossum ringspot virus(ORSV) are the two main viruses which often infect Phalaenopsis severely and lead to huge economic losses to the industry. In presented paper, these two viruses were detected rapidly, which would provide a basis for breeding Phalaenopsis of resistance to both viruses of CyMV and ORSV. 【Method】The specific primers were designed based on CyMV coat protein(CP) gene and ORSV CP gene sequence which had been reported in NCBI, and antisense RNA technology was used in this study. 【Result】12 samples of Phalaenopsis by using designed specific primers were detected, and 25% of them contained the two viruses, and the cloning products were clear and no redundancy from the same PCR reaction. 125 bp fragment of CyMV CP gene and 164 bp fragment of ORSV CP gene from the leaf tissue of infected Phalaenopsis were obtained. The CP gene fragment of CyMV linked to the expression vector p CAMBIA1300 in 3'→5'orientation, and named p CAMBIA1300-CyMV which could obtain 125 bp fragment by PCR and restriction enzyme digestion. The CP gene fragment of ORSV linked to the constructed vector p CAMBIA1300-CyMV in 3'→5'orientation, and designated p CAMBIA1300-CyMV-ORSV which could obtain 289 bp fragment by PCR and restriction enzyme digestion. 【Conclusion】CyMV and ORSV by using the specific primers could be detected simultaneously in the same PCR reaction system, and the double-antisense expression vector containing two kinds of the virus CP genes was constructed successfully. [ABSTRACT FROM AUTHOR]