Molecular, ecological, and transgenic studies of cucumber mosaic virus and its satellite RNA.

摘要

Cucumber mosaic virus (CMV) is a single-stranded, tripartite (+) RNA virus which causes many important plant diseases worldwide. It sometimes harbors a satellite RNA of ca. 340 bases which can greatly affect symptomatology, and has been responsible for one severe and two minor disease occurrences in the field. In my work, I have cloned the satellite of the white leaf strain of CMV and have obtained in vitro transcripts. These transcripts induce the same symptoms as native satellite in tomato and tobacco when co-inoculated with CMV genomic RNA. The symptom-inducing activity greatly decreases with the number of non-satellite bases at the 5{dollar}spprime{dollar} end of the transcript. I have developed a means of cloning CMV satellites so as to obtain highly active transcripts with a minimum of manipulation. Dimeric transcripts were not more active than monomers; satellite cDNA was inactive. Next investigated was the occurrence of CMV satellites in nature. Analysis of 134 CMV-infected field field samples from Bermuda and western New York State by dot blot and northern hybridization yielded only two samples containing CMV satellite. Using ELISA (Enzyme-Linked Immunosorbent Assay) with antisera representing two CMV serogroups, the samples divided fairly well into serogroups, with both groups well represented. A severe tomato disease from China was also analyzed. It was determined to be caused by a severe strain of CMV, which induced severe symptoms in many hosts. A satellite that induces necrosis in tomato was found associated with the isolate, but most of the severity seen in the field could be attributed to the viral genomic RNA. Finally, the beginnings of a project for the insertion of satellite cDNA into the tobacco genome are described and future experiments with the transgenic plants are outlined. In the final chapter of the thesis, I discuss the different activities of the various types of in vitro transcripts and what this implies about satellite replication. In addition, the rarity of CMV satellites in the field may have implications for the use of transgenic satellite as a source of resistance in field crops.