Use of Hairpin RNA Constructs for Engineering Plant Virus Resistance

Genetically engineered resistance to protect plants against virus infections can be based on protein- and RNA-mediated defense mechanisms. RNA silencing that leads to high-level virus resistance is triggered by virus-specific double-stranded (ds)RNA. The most efficient means to produce such dsRNA in transgenic plants is the introduction and expression of hairpin (hp) RNA constructs. Successful induction of the RNA silencing pathway is witnessed by the accumulation of virus-specific small interfering (si)RNAs that guide destruction of complementary viral RNA. Here, we describe strategies and methods for the efficient generation of hpRNA constructs and for the extraction and detection of siRNAs.