title: Using RNA-mediated interference feeding strategy to screen for genes involved in body size regulation in the nematode C. elegans.
abstract: Double-strand RNA-mediated interference (RNAi) is an effective strategy to knock down target gene expression. It has been applied to many model systems including plants, invertebrates and vertebrates. There are various methods to achieve RNAi in vivo. For example, the target gene may be transformed into an RNAi vector, and then either permanently or transiently transformed into cell lines or primary cells to achieve gene knockdown effects; alternatively synthesized double-strand oligonucleotides from specific target genes (RNAi oligos) may be transiently transformed into cell lines or primary cells to silence target genes; or synthesized double-strand RNA molecules may be microinjected into an organism. Since the nematode C. elegans uses bacteria as a food source, feeding the animals with bacteria expressing double-strand RNA against target genes provides a viable strategy. Here we present an RNAi feeding method to score body size phenotype. Body size in C. elegans is regulated primarily by the TGF- β-llike ligand DBL-1, so this assay is appropriate for identification of TGF-β signaling components. We used different strains including two RNAi hypersensitive strains to repeat the RNAi feeding experiments. Our results showed that rrf-3 strain gave us the best expected RNAi phenotype. The method is easy to perform, reproducible, and easily quantified. Furthermore, our protocol minimizes the use of specialized equipment, so it is suitable for smaller laboratories or those at predominantly undergraduate institutions.
authors: Department of Science, Borough of Manhattan Community College, City University of New York, NY, USA. jrice@bmcc.cuny.edu
authors: Jun
authors: J
authors: Liang
authors: Sheng
authors: S
authors: Xiong
authors: Cathy
authors: C
authors: Savage-Dunn
mesh_terms: D000818:Animals; D049628:Body Size / Q000235:genetics; D017173:Caenorhabditis elegans / Q000033:anatomy & histology* / Q000235:genetics*; D055785:Gene Knockdown Techniques; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000235:genetics; D012324:RNA-Dependent RNA Polymerase / Q000235:genetics; D016212:Transforming Growth Factor beta / Q000235:genetics
chemical_list: D012330:RNA, Double-Stranded; D016212:Transforming Growth Factor beta; D012324:RNA-Dependent RNA Polymerase; C472756:RNA-directed RNA polymerase RRF-3, C elegans