Digital PCR Genotyping of Pepino Mosaic Virus.

Pepino mosaic virus (PepMV) is a plant pathogen causing significant economic losses in tomato production. Sensitive, reliable, and robust detection methods are crucial for containing the spread of PepMV and reducing its damaging effects. Digital PCR (dPCR) presents several advantages to conventional real-time quantitative PCR (qPCR), including absolute quantification ability, robust quantitative multiplexing capabilities, and straightforward result analysis. Furthermore, dPCR is especially suitable for analysis of complex samples due to its remarkable tolerance to PCR inhibitors, which makes it a promising method for plant virus genotyping. In this chapter, we present two protocols for PepMV genotyping and quantification using one-step reverse transcription digital PCR (RT-dPCR). The first protocol outlines four simplex assays, while the second describes two duplex assays for precise and comprehensive genotyping of PepMV variants.