Genome-wide RNAi of C. elegans using the hypersensitive rrf-3 strain reveals novel gene functions.
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.
Simmer
Femke
F
Hubrecht Laboratory, Centre for Biomedical Genetics, Utrecht, The Netherlands.
Moorman
Celine
C
van der Linden
Alexander M
AM
Kuijk
Ewart
E
van den Berghe
Peter V E
PV
Kamath
Ravi S
RS
Fraser
Andrew G
AG
Ahringer
Julie
J
Plasterk
Ronald H A
RH
D000818:Animals; D017173:Caenorhabditis elegans / Q000235:genetics*; D029742:Caenorhabditis elegans Proteins / Q000378:metabolism; D002874:Chromosome Mapping; D002875:Chromosomes / Q000648:ultrastructure; D003001:Cloning, Molecular; D005786:Gene Expression Regulation; D015723:Gene Library; D017238:Genes, Helminth; D005821:Genetic Techniques; D016678:Genome; D023281:Genomics; D008954:Models, Biological; D009154:Mutation; D010641:Phenotype; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000737:chemistry; D013045:Species Specificity
D029742:Caenorhabditis elegans Proteins; D012330:RNA, Double-Stranded