Detection and Characterization of Subgenomic RNA in Plant Viruses

Most plant viruses contain single-stranded RNA genomes and many also give rise to positive-strand subgenomic RNAs. Subgenomic RNAs are defined as RNAs related by sequence to a viral genome, which are generated during the course of infection. These subgenomic RNAs are often found encapsidated, the presence of some being more obvious than that of others. In many cases, however, certain subgenomic RNAs are found only in infected plant cells, not in virions. A number of new biochemical techniques have been developed or modified that enable subgenomic RNAs to be detected, isolated, purified, and characterized. These new procedures are never static. They are forever being modified, naturally, always for the better. This chapter provides an overview of a range of techniques used to detect and characterize subgenomic RNAs. It describes techniques suited for analyzing positive-strand RNA plant viruses; however, many of the same techniques described here can be used for or characterizing subgenomic RNAs of either negative-strand RNA viruses, double-stranded RNA viruses, or DNA viruses. The ends of RNA molecules can be determined by the use of techniques described here. The techniques provide a better understanding of the relationship of the components involved in the replication and spread of RNA plant viruses. With the aid of the techniques of molecular cloning and the expression of cloned genes, sufficient amounts of virus-encoded proteins should become available for study of their function and involvement in virus replication, spread, and pathology.