title: Dissecting systemic RNA interference in the red flour beetle Tribolium castaneum: parameters affecting the efficiency of RNAi.
abstract: The phenomenon of RNAi, in which the introduction of dsRNA into a cell triggers the destruction of the corresponding mRNA resulting in a gene silencing effect, is conserved across a wide array of plant and animal phyla. However, the mechanism by which the dsRNA enters a cell, allowing the RNAi effect to occur throughout a multicellular organism (systemic RNAi), has only been studied extensively in certain plants and the nematode Caenorhabditis elegans. In recent years, RNAi has become a popular reverse genetic technique for gene silencing in many organisms. Although many RNAi techniques in non-traditional model organisms rely on the systemic nature of RNAi, little has been done to analyze the parameters required to obtain a robust systemic RNAi response. The data provided here show that the concentration and length of dsRNA have profound effects on the efficacy of the RNAi response both in regard to initial efficiency and duration of the effect in Tribolium castaneum. In addition, our analyses using a series of short dsRNAs and chimeric dsRNA provide evidence that dsRNA cellular uptake (and not the RNAi response itself) is the major step affected by dsRNA size in Tribolium. We also demonstrate that competitive inhibition of dsRNA can occur when multiple dsRNAs are injected together, influencing the effectiveness of RNAi. These data provide specific information essential to the design and implementation of RNAi based studies, and may provide insight into the molecular basis of the systemic RNAi response in insects.
authors.affiliation: Division of Biology, Kansas State University, Manhattan, Kansas, United States of America.
authors.forename: Sherry C
authors.initials: SC
authors.lastname: Miller
authors.forename: Keita
authors.initials: K
authors.lastname: Miyata
authors.forename: Susan J
authors.initials: SJ
authors.lastname: Brown
authors.forename: Yoshinori
authors.initials: Y
authors.lastname: Tomoyasu
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D017931:DNA Primers / Q000235:genetics; D020868:Gene Silencing; D005821:Genetic Techniques; D049452:Green Fluorescent Proteins / Q000378:metabolism; D008957:Models, Genetic; D016133:Polymerase Chain Reaction / Q000379:methods; D012313:RNA / Q000032:analysis*; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000032:analysis; D014231:Tribolium / Q000235:genetics* / Q000502:physiology
chemical_list: D017931:DNA Primers; D012330:RNA, Double-Stranded; D049452:Green Fluorescent Proteins; D012313:RNA