Whole-animal high-throughput screens: the C. elegans model.
The nematode Caenorhabditis elegans shows a high degree of conservation of molecular pathways related to human disease, yet is only 1-mm long and can be considered as a microorganism. Because of the development of a simple but systematic RNA-interference (RNAi) methodology, C. elegans is the only metazoan in which the impact of "knocking-down" nearly every gene in the genome can be analyzed in a whole living animal. Both functional genomic studies and chemical screens can be carried out using C. elegans in vivo screens in a context that preserves intact cell-to-cell communication, neuroendocrine signaling, and every aspect of the animal's metabolism necessary to survive and reproduce in lab conditions. This feature enables studies that are impossible to undertake in cell-culture-based screens. Although genome-wide RNAi screens and limited small-molecule screens have been successfully performed in C. elegans, they are typically extremely labor-intensive. Furthermore, technical limitations have precluded quantitative measurements and time-resolved analyses.In this chapter, we provide detailed protocols to carry out automated high-throughput whole-animal RNAi and chemical screens. We describe methods to perform screens in solid and liquid media, in 96 and 384-well format, respectively. We describe the use of automated handling, sorting, and microscopy of worms. Finally, we give information about worm-adapted image analysis tools to quantify phenotypes. The technology presented here facilitates large-scale C. elegans genetic and chemical screens and it is expected to help shed light on relevant biological areas.
O'Rourke
Eyleen J
EJ
Department of Molecular Biology, Massachusetts General Hospital, Department of Genetics, Harvard Medical School, Boston, MA, USA. eorourke@molbio.mgh.harvard.edu
Conery
Annie L
AL
Moy
Terence I
TI
D000818:Animals; D030801:Animals, Genetically Modified; D017173:Caenorhabditis elegans / Q000235:genetics* / Q000254:growth & development*; D015723:Gene Library; D034622:RNA Interference / Q000502:physiology*; D015996:Survival Rate