Effectiveness of specific RNA-mediated interference through ingested double-stranded RNA in Caenorhabditis elegans

By R. Kamath, Maruxa Martínez‐Campos, Peder Zipperlen, Andrew G. Fraser, J. Ahringer. Genome Biology, 2000

1,378 citations recorded by the source corpus.

BackgroundIn Caenorhabditis elegans, injection of double-stranded RNA (dsRNA) results in the specific inactivation of genes containing homologous sequences, a technique termed RNA-mediated interference (RNAi). It has previously been shown that RNAi can also be achieved by feeding worms Escherichia coli expressing dsRNA corresponding to a specific gene; this mode of dsRNA introduction is conventionally considered to be less efficient than direct injection, however, and has therefore seen limited use, even though it is considerably less labor-intensive.ResultsHere we present an optimized feeding method that results in phenotypes at least as strong as those produced by direct injection of dsRNA for embryonic lethal genes, and stronger for genes with post-embryonic phenotypes. In addition, the interference effect generated by feeding can be titrated to uncover a series of hypomorphic phenotypes informative about the functions of a given gene. Using this method, we screened 86 random genes on consecutive cosmids and identified functions for 13 new genes. These included two genes producing an uncoordinated phenotype (a previously uncharacterized POU homeodomain gene, ceh-6, and a gene encoding a MADS-box protein) and one gene encoding a novel protein that results in a high-incidence-of-males phenotype.ConclusionsRNAi by feeding can provide significant information about the functions of an individual gene beyond that provided by injection. Moreover, it can be used for special applications for which injection or the use of mutants is sometimes impracticable (for example, titration, biochemistry and large-scale screening). Thus, RNAi by feeding should make possible new experimental approaches for the use of genomic sequence information.

Subfields: Aging, Molecular Biology

View Paper: https://www.semanticscholar.org/paper/49961bf62af11cd7bea8c91034bb930cdf5e8e29

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Citations in the corpus

Loss of the putative RNA-directed RNA polymerase RRF-3 makes C. elegans hypersensitive to RNAi. (2002, methodology): “Biology Graduate Program (pk1426) and wild-type animals were fed as described Johns Hopkins University by Kamath et al. [ 5 ].”
Direct inhibition of the longevity-promoting factor SKN-1 by insulin-like signaling in C. elegans. (2008, methodology): “RNAi Feeding RNAi was carried out as previously described, with empty pL4440 as the control ((Kamath et al., 2001); supplementary methods).”
Genome-Wide RNAi of C. elegans Using the Hypersensitive rrf-3 Strain Reveals Novel Gene Functions (2003, methodology): “Other global RNAi screens have been recently performed in C. elegans using this RNAi library or other techniques (Gönczy et al. 2000; Maeda et al. 2001; Dillin et al. 2002; Piano et al. 2002; Ashrafi et al. 2003; Lee et al. 2003; Pothof et al. 2003).”
The TOR pathway interacts with the insulin signaling pathway to regulate C. elegans larval development, metabolism and life span (2004, background): “In C. elegans, insulin/IGF signaling regulates larval development and adult life span (Kenyon et al., 1993; Kimura et al., 1997).”