A Single-Step Multiplex RT-PCR to Detect Enteric Viruses

A Single-Step Multiplex RT-PCR to Detect Enteric Viruses

  A single-step multiplex reverse transcription-polymerase chain reaction (RT-PCR) assay that detects and identifies Adenovirus, Astrovirus, Norovirus genogroup I (GI) and II (GII), and Rotavirus in stool samples is described. The primers for Adenovirus, Norovirus GI and GII, and Rotavirus were designed from the capsid region of target viral genome and one primer set for Astrovirus was designed from the RNA-dependent RNA polymerase gene. The multiplex RT-PCR reaction mixture containing five sets of primers amplified each expected product independently and specifically in a single-step with the One Step RNA PCR Kit (AMV) (Takara). The multiplex RT-PCR was validated against simplex RT-PCR for Astrovirus, Norovirus GI, Norovirus GII, and Rotavirus, and against simplex PCR for Adenovirus. The multiplex RT-PCR was equally specific in detecting Adenovirus, Astrovirus, Norovirus GI and GII, and Rotavirus compared with simplex RT-PCR and PCR.