黄瓜花叶病毒2b 蛋白关键氨基酸的致病性分析

MV recombinant FRad352b-CMV is generated by replacing 2b gene in Fny-CMV with 2b gene of Rad35-CMV, inducing mild mosaic. In this study, we found FRad352b-CMV severe mosaic symptom mutation of FRad35Leu2b-CMV. Northern blotting and sequence analysis showed: the Leu55 of FRad352bLeu-CMV 2b protein plays an important role in the pathogenicity mediated by the 2b protein. we inoculated FRad35Leu2b-CMV and FRad352b-CMV on GFP-transgenic N. benthamiana plant (line 16c), in which GFP was pre-silenced by by infiltration with a hypervirulent strain of A.tumefaciens carrying the 35S-GFP plasmid. It is concluded that Leu55 plays a critical role in suppressing PTGS. we constructed agrobacteria transient expression vectors of 2b gene of FRad35Leu2b-CMV, FRad35Leu2b-CMV, 35S-2bPro and 35S-2bLeu. When agrobacteria carrying these vectors co-infiltrated with that carrying 35S-GFP on Nicotiana benthamiana plant (line 16c), northern blot analyzes accumulation of GFP mRNA and GFP siRNA in the tested plants at 3d丆7d丆and 14d. We speculate that high virulence mediated by 2bLeu is achieved through blocking systemic movement of viral siRNA, making CMV systemically move and replicate more effectively, then resulting to induction of severe viral symptom on hosts.