Gene Expression Analysis Of Psuedo-recombination Between Cucumber Mosaic Virus M And Fny Strains In Systemically Infected Nicotiana Benthamiana Seedlings

Cucumber mosaic virus (CMV) has a worldwide distribution and a very wide host range. In fact it has the reputation of having the widest host range of any known plant virus. In this study,we researched pathogenic mechanismon and recovery mechanismon of CMV technologys of infectious clone,virus recombination and real-time fluorescence PCR.Then, the pathogenic mechanism of CMV-M and CMV-Fny, and the causes of recovery phenomenon were preliminarily explored.Using reverse transcription-PCR (RT-PCR), full-length cDNA clones of Cucumber mosaic virus M strain (CMV-M) were constructed. Though pseudo-recombination between CMV-M (RNA2 and RNA3) and CMV-Fny genomic RNAs, the pseudo-recombinants F1M2F3, F1F2M3 and F1M2M3 were successful obtained. The seedings of Nicotiana tabacum cv. White Burley inoculated with pseudo-recombinants F1M2F3, F1F2M3 and F1M2M3 produced necrosis, chlorosis, vein clearing, labefaction, linearization of leaf apex, etc. Based on comparison of symtoms between pseudo-recombinants and wild type virus, the key virulent factors were speculated. The key factor which induced mosaic was CP, while RNA2 of CMV-M and CMV-Fny was involved with necrosis and linearization of leaf apex respectively. Real-time fluorescence quantitative PCR analysis indicated that symptom difference in N. tabacum cv. White Burley induced by CMV-M, CMV-Fny, F1M2F3, F1F2M3 and F1M2M3 was not related to the accumulation level of CMV genomic RNAs.Primers and probes were designed for detection of CMV RNA1, RNA2, RNA3, RNA4. Real-time fluorescence quantitative PCR analysis reflected that the loading of CMV-M RNA1, RNA2, RNA3 in the disease and recovey leaves shows no difference,and maintained at comparatively high levels.Howerver, a sudden decrease of CMV RNA4 happened in the recovey leaves. CMV-M RNAs were severely degraded at 30dpi. Degree of degradation in recovey leaves was most serious, the loading of RNA1,RNA2,RNA3,RNA4 had decreased 36,139,100,320 times, separately. We concluded that there is some relationship between RNA4 and recovery mechanismon, and the degradation mechanismon is more powerful in recovey leaves than other leaves.