Infection Of Lilies And Daffodils Part Of The Linear Plant Virus Coat Protein Gene, Prokaryotic Expression, Antiserum Preparation And Detection Applications

Virus diseases of narcissus(Narcissus spp.) and lily(Lilium spp.) are widespread throughout the world, These viruses accumulate in bulbs because of the vegetative propagation of the crop. Narcissus and lily are economically important crops in China, and filamentous virus diseases cause severe losses of yield and poor bulb quality. Narcissus yellow stripe virus (NYSV),Narcissus mosaic virus(NMV), Narcissus latent virus(NLV), Narcissus common latent virus(NCLV), Narcissus degeneration virus, (NDV), Ornithogalum mosaic virus(OrMV) and Narcissus late season yellows virus(NLSYW) are the most common filamentous viruses in narcissus, Lily mottle virus (LMoV), Lily virus X(LVX) and Lily symptomless virus (LSV) are same in lily. Because these viruses often infect the bulbs in mixture and are difficult to separate,therefore,they are not easy to purify for production of high quality antisera for studying of serological relationship among them,which will be helpful for virus identification and detection.The specific primers were designed according to the published viral sequences, to amplify the coat protein genes of NYSV, NCLV, OrMV, LSV, LVX and LMoV respectively. The CP gene of those viruses were obtained by PCR and RT-PCR. The plasmids(pGEM-T) contain these CP genes were constructed and the CP genes were sequenced and confirmed to be correct. Then the CP genes were cloned into pSBET plasmids and prokaryotic expressed in E.coli BL21(DE3)plysS. The antisera were raised in mouse which were spectific to their homologous CPs,western-blot analysis indicated that there was no detectable reactions among the narcissus viruses (NYSV NCLV and OrMV) and also among the lily viruses (LMoV, LVX and LSV). Western-blot analysis of NYSV, NCLV, OrMV, LSV, LVX and LMoV with some other virus which were in the same genus indicated that there were detectable reactions, although usually weak, among NYSV and ScaMV, TuMV, ZYMVandDsMV.There were no detectable reactions between antisera from OrMV, NCLV, LMoV, LSV and LVX and CPs from some other virus which were in the same genus with OrMV, NCLV, LMoV, LSV and LVX.ELISA detections with the field samples showed that NYSV, NCLV and OrMV accured in narcissus plants in China, and LMoV, LSV in lily plants in Zhejiang province. LVX was not detected.