Molecular Cloning and Complete Nucleotide Sequence of Rice Ragged Stunt Oryzavirus Genome Segment 10

Abstract

cDNAs to rice ragged stunt Oryzavirus genome segment 10 (S10) were made by oligo-dT primed 1st strand cDNA synthesis after poly A addition to 3' terminal of viral transcripts and cloned in a bacterial plasmid pBR322. The overlapping cDNA clones were sequenced and the assembled sequence covered the full-length of the genome except for 22 nucleotides from the 5' terminus, which were determined by a direct RNA sequencing method. RRSV S10 is 1162 nucleotide long and an inverted repeat structure containing 6 nucleotides was found in the terminal region. It has a mini ORF in addition to the down stream long ORF. The upstream mini ORF encodes 12 amino acids. Presence of this mini ORF was confirmed by sequencing another full-length cDNA clones generated by reverse transcription coupled with a polymerase chain reaction. The long ORF encodes a polypeptide of 297 amino acids (M_r 32.4K).