Tools and techniques for production of double-stranded RNA and its application for management of plant viral diseases
Andreas E. Voloudakis, Maria C. Holeva, Athanasios Kaldis and Dongho Kim
from: Genes, Genetics and Transgenics for Virus Resistance in Plants (Edited by: Basavaprabhu L. Patil). Caister Academic Press, U.K. (2018) Pages: 119-140.
Due to the rapidly growing global population, food production and security is the major challenge of agriculture. Plant viruses are obligate parasites that in some instances could cause up to 100% losses in a crop (e.g. maize streak disease). Although difficult to accurately determine the global economic impact that plant viruses have on agriculture, it is estimated that US$ 60 billion loss in crop yields worldwide each year is due to plant viral diseases. RNA silencing (RNA interference, RNAi) is a conserved endogenous pathway of all higher eukaryotes, which controls gene expression. RNAi is induced by double-stranded RNA (dsRNA) and allows the cell to recognize aberrant genetic material in a highly sequence-specific manner ultimately leading to its degradation, thus protecting the cell from subcellular pathogens, such as viruses and transposons. DsRNA-mediated resistance has been exploited in transgenic plants to convey resistance against viruses and against insects, vectors of plant viruses, via host induced gene silencing (HIGS). A non-transgenic approach employing RNAi has been used where enzymatically synthesized specific dsRNA molecules, when applied directly onto plant tissue, induce resistance against the cognate virus; as a result dsRNA molecules could be efficacious antiviral agents for crop protection. Next generation sequencing and bioinformatics analyses have provided a plethora of information and useful tools for the design and study of dsRNA application. In this chapter, the different methods for dsRNA production, both in vitro and in vivo, the means of direct application of the dsRNA molecules onto plants and several examples of non-transgenic dsRNA-mediated resistance are presented read more ...