We have recently completed cloning and sequencing the entire genome of dengue type 4 virus. The RNA genome of dengue type 4 virus is 10,644 nucleotides long and contains a single long open reading frame (ORF) encoding a polyprotein chain of 3386 amino acids. The positive strand RNA of dengue 4 serves as a messenger RNA for the synthesis of a polyprotein chain which is post-translationally cleaved into the full complement of virus proteins. We have initiated the expression of single or multiple dengue type 4 virus proteins from our cDNA clones using baculovirus as a vector. A DNA fragment encoding: (1) the C, M, and E structural proteins, (b) the NS1 nonstructural protein or (c) the NS5 nonstructural protein has been inserted into a baculovirus intermediate vector. Subsequently baculovirus-dengue recombinants were identified and purified by plaque isolation. Currently we are screening these baculovirus recombinants for the expression of dengue proteins. Expressed proteins will be used to immunize animals for preparation of antisera and these antisera will in turn be used for identification of dengue viral proteins produced during infection. Some of the expressed proteins may also prove useful as immunogens for use in the immunoprophylaxis of dengue virus infection and disease.
