Molecular detection and identification of potato isolates of Tobacco rattle virus

Abstract

Sixteen isolates of Tobacco rattle virus (TRV) obtained from the United States and Canada were characterized in the present study. Nucleotide-sequence analysis of open reading frame 4 (ORF4) in RNA1 of seven test isolates revealed a high degree of genetic diversity. Potato isolates of TRV clustered in several distinct groups, with isolates from the same geographic region falling into different groups. Phylogenetic analysis of TRV isolates based on nucleotide sequences and on restriction fragment length polymorphism (RFLP) produced similar groupings. TRV was detected in various potato tissues of inoculated, field-grown plants, including progeny tubers, leaves, and roots, by reverse transcription – polymerase chain reaction (RT–PCR), using a pair of TRV-specific primers targeting ORF4. TRV RNA was detected in highly diluted total RNA preparations (1 / 15 625) and in composite tuber samples (200–400 tubers). RT–PCR followed by RFLP analysis was shown to be an efficient procedure for specific and sensitive detection and identification of TRV in potato.

Keywords:

• Tobravirus
• Tobacco rattle virus
• potato isolates
• 16 kDa protein gene
• phylogenetic analysis
• molecular detection