The analysis of cloned rotavirus genes will greatly enchance our knowledge of the molecular biology of rotavirus. In addition to identification of important regulatory signals (promoters, ribosomal binding sites, etc.), analysis of cloned cDNA can yield information concerning amino acid sequences of structural proteins. Expression of rotavirus antigens by insertion of cloned DNA from VP7 or VP3 into various vectors may provide us with safer RV vaccines. Although rotavirus cDNA clones representing VP7 have been inserted into bacterial and vaccinia expression vectors, the proteins expressed have not been highly immunogenic. It is possible, however, that insertion of the VP3 protein gene into such vectors will result in the expression of a more antigenic protein. VP3 and VP7 clones are being engineered for insertion into adenovirus and bacterial expression vectors which are currently under development. We are currently attempting to transfect rotavirus ss RNA from a given serotype into cells infected with a rotavirus strain of a different serotype in an attempt to recover (under selective pressure) rotavirus reassortants containing the transfected gene. If these experiments yield positive results it will be possible to attempt the rescue of rotavirus modified by in vitro mutagenesis of cloned DNA. Success in developing this methodology will also allow us to probe the function of certain areas of the rotavirus genome (i.e. serotype specific regions in the VP7 gene, cleavage region in the fourth gene, etc.) and could be of considerable help in constructing cross-reactive or further attenuated rotavirus vaccine candidates.
