cDNA cloning of each genomic segment of the group B rotavirus ADRV: molecular characterization of the 11th RNA segment

Abstract

The group B noncultivatable rotavirus, ADRV, was purified from infected stool specimens. Double-stranded RNA was extracted, polyadenylated, reverse transcribed into cDNA, and cloned into plasmid vector pAT153. Each cDNA clone hybridized to a single ADRV RNA segment and cDNA clones of each genomic RNA segment were identified. Sequencing of genomic RNA and a full-length cDNA clone (AD63) determined that ADRV RNA segment 11 is 631 bases in length and contains a single open reading frame of 170 amino acids with a calculated molecular weight of 19.9 kDa and a pl of 6.2. The RNA 5'- and 3'-termini contain untranslated regions of 58 and 63 bases, respectively, and are complementary to each other. A comparison of encoded ADRV gene 11 amino acids with the NS26 protein of group A rotaviruses demonstrates a distant similarity between the two proteins. Further analysis and use of these ADRV clones should aid in determining the gene coding assignments for group B proteins as well as for diagnostic detection of group B or ADRV-specific nucleic acids in specimens.