A fitness assay for comparing RNAi effects across multiple C. elegans genotypes Mark Elvin¹, Basten L Snoek², Martin Frejno¹, Ulrike Klemstein¹, Jan E Kammenga², Gino B Poulin¹ 1University of Manchester, Faculty of Life Sciences, Manchester, M13 9PT, United Kingdom, 2Wageningen University, Nematology, Wageningen, 7608PB, Netherlands RNAi technology by feeding C. elegans with E. coli containing dsRNA has significantly contributed to increase our understanding of many different fields of biology, including genetics, molecular biology, developmental biology and functional genomics. Most of this research has been carried out in a single genotype or genetic background. However, RNAi effects in one genotype do not reveal the allelic effects that segregate in natural populations and contribute to phenotypic variation. Here we present a method that allows for rapidly comparing RNAi effects among diverse genotypes at an improved high throughput rate. It is based on assessing the fitness of a population of worms by measuring the rate at which E. coli is consumed, the feeding curve. Critically, we demonstrate the analytical power of this method by using the feeding curves of worms treated with RNAi for QTL mapping. We tested a panel of 12 genes (tag-214, mel-26, rab-5, pos-1, par-6, gld-1, let-502, lin-31, par-1, smo-1 and mpk-1) on a recombinant inbred population derived from a cross between Bristol and a natural isolate from Hawaii. Hawaii has lost RNAi sensitivity in the germline. We confirmed that polymorphisms in ppw-1 contribute to this loss of RNAi sensitivity as for many RNAi treatments the QTLs are found at the ppw-1 locus, but moreover we found that other loci are also likely to be important. In summary, we have established a fast method that improves the throughput of RNAi in liquid, that generates quantitative data, that is easy to implement in most laboratories, and importantly that enables QTL mapping using RNAi. Funding: the EU 7th Framework Programme under the Research Project PANACEA, contract nr 222936.
|Title of host publication||Abstracts of papers presented at the Evolution of Caenorhabditis and Other Nematodes, Cold Spring Harbor Laboratory, New York, USA, 3-6 April 2012|
|Publication status||Published - 2012|
|Event||Evolution of Caenorhabditis and Other Nematodes, Cold Spring Harbor Laboratory, New York, USA - |
Duration: 3 Apr 2012 → 6 Apr 2012
|Conference||Evolution of Caenorhabditis and Other Nematodes, Cold Spring Harbor Laboratory, New York, USA|
|Period||3/04/12 → 6/04/12|
Elvin, M., Snoek, L. B., Frejno, M., Klemstein, U., Kammenga, J. E., & Poulin, G. (2012). A fitness assay for comparing RNAi effects across multiple C. elegans genotypes. In Abstracts of papers presented at the Evolution of Caenorhabditis and Other Nematodes, Cold Spring Harbor Laboratory, New York, USA, 3-6 April 2012 (pp. 46-46)