Subcellular Colocalization Assay of Host Factors with Viral Replication Complex in the dsRNA Reporter Nicotiana benthamiana.

ABSTRACT

As obligate pathogens, plant viruses co-opt several host factors for viral replication. Double-stranded RNA (dsRNA) plays important roles in plants, including eliciting innate immune responses and RNA interference. dsRNA also represents the genetic entities of a number of viruses and is a marker of infection by positive-sense single-stranded RNA viruses. Previous detection methods for RNA viruses basically relied on immunological methods, but later researchers discovered that the dsRNA-binding domain of the Flock house virus B2 protein is a perfect alternative to the J2 mAb for sensitive and rapid detection of long dsRNA in vitro and in vivo, and developed B2GFP transgenic Nicotiana benthamiana line. This method describes in detail how to visualize host factors in the viral replication complex in time and space with the help of B2GFP transgenic plants, exemplified by Turnip mosaic virus (TuMV), a representative virus member of the Potyviruses.