Genome-wide RNAi of C. elegans using the hypersensitive rrf-3 strain reveals novel gene functions - PubMed (original) (raw)

Genome-wide RNAi of C. elegans using the hypersensitive rrf-3 strain reveals novel gene functions

Femke Simmer et al. PLoS Biol. 2003 Oct.

Abstract

RNA-mediated interference (RNAi) is a method to inhibit gene function by introduction of double-stranded RNA (dsRNA). Recently, an RNAi library was constructed that consists of bacterial clones expressing dsRNA, corresponding to nearly 90% of the 19,427 predicted genes of C. elegans. Feeding of this RNAi library to the standard wild-type laboratory strain Bristol N2 detected phenotypes for approximately 10% of the corresponding genes. To increase the number of genes for which a loss-of-function phenotype can be detected, we undertook a genome-wide RNAi screen using the rrf-3 mutant strain, which we found to be hypersensitive to RNAi. Feeding of the RNAi library to rrf-3 mutants resulted in additional loss-of-function phenotypes for 393 genes, increasing the number of genes with a phenotype by 23%. These additional phenotypes are distributed over different phenotypic classes. We also studied interexperimental variability in RNAi results and found persistent levels of false negatives. In addition, we used the RNAi phenotypes obtained with the genome-wide screens to systematically clone seven existing genetic mutants with visible phenotypes. The genome-wide RNAi screen using rrf-3 significantly increased the functional data on the C. elegans genome. The resulting dataset will be valuable in conjunction with other functional genomics approaches, as well as in other model organisms.

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Conflict of interest statement

The authors have declared that no conflicts of interest exist.

Figures

Figure 1

Figure 1. Comparison of Different RNAi Experiments of Chromosome I Using Wild-Type Bristol N2 and rrf-3

Differences between different laboratories or investigators and between experiments done within the same laboratory and by the same investigators are observed. Ovals represent the amount of bacterial clones that gave an RNAi phenotype in an experiment. Areas that overlap represent clones for which in both experiments an RNAi phenotype was detected. Differences and overlap between an RNAi experiment done with the rrf-3 mutant strain and the data obtained by Fraser et al. (2000) done with the standard laboratory strain, Bristol N2 (A); N2 and rrf-3 tested at the same time within our laboratory (B); experiments done with N2 in two different laboratories: this study (‘NL') and Fraser et al. (2000) (C); two experiments done with the same strain, rrf-3, within our laboratory (D).

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