Project Leader:   
Don Moerman

Involved Institution:  
University of British Columbia  

Technology Applications:   
Gene knockout tools; Gene function models; Developmental models

Research Funding Program:   
Competition I

Humans share approximately 7000 genes with Caenorhabditis elegans, a transparent worm used by geneticists as a model organism to study gene function. The shared genes, when dysfunctional in humans, lead to inherited diseases, susceptibility to cancers, or other health concerns. Therefore, understanding the basic biological function of these genes in a model organism like C. elegans could lead to changes in human medical care. To identify a gene’s role in development, the gene is inactivated or ‘knocked out’ and this research group, in collaboration with the Oklahoma Medical Research Foundation, has improved the technology used to create the knockouts. The project plans to obtain targeted mutations in 2000 of the C. elegans genes shared with humans. Researchers around the world can then use those gene knockouts for further study. In the first two years, this project has identified mutations in over 490 genes and nearly doubled the number of genes with mutations available to the C. elegans research community.

The nematode C. elegans is a powerful research tool to investigate fundamental questions about biology. First of all, it was the first multicellular organism for which a complete DNA sequence was known. It is also the only multicellular organism that has a complete cell lineage map beginning from a single cell embryo to the adult. Plus, complex cellular and developmental processes can be examined because individual cells can be observed.

The C. elegans research community is pursuing the goal of complete, integrated molecular understanding of C. elegans development and physiology, which requires exceptional cooperation among researchers. To tackle this large project of targeted mutations, an international consortium has been formed, including the Oklahoma Medical Research Foundation, Genome BC’s investigators at the University of British Columbia and the Genome Sciences Centre at the BC Cancer Agency.

The C. elegans 97MB genome encodes approximately 19,000 predicted proteins. Using traditional genetic approaches, about 1,500 C. elegans genes were identified with mutant phenotypes. To identify the remaining 17,000 genes, researchers are using a reverse genetics approach to create mutations. Developed by Dr. Barstead at the Oklahoma Medical Research Foundation, the PCR-based technique is related to transposable elements but substitutes a chemical mutagen for Tc1. The choice of genes targeted for knockout is decided in consultation with the scientific community; there is a website where researchers worldwide can make requests for a gene with a human homolog to be targeted for deletion.

Over the past two years, this project has identified deletion mutations in over 490 genes, five times the number reported by any other group. Plus, analysis has been completed on 381 of these knockout mutations, meaning that homozygous strains have been made for those that are viable as homozygotes. For those that are homozygous inviable, balanced heterozygotes have been constructed. The mutations have also been sequenced, with the breakpoints determined and the data submitted to WormBase, the main database for C. elegans information. Furthermore, all deletion-containing lines are sent to the Caenorhabditis Genetics Center in St. Paul, Minnesota, providing the knockouts to the larger research community for functional analysis.

Project publications:
Edgely, M., D'Souza, A., Moulder, G., McKay, S., Shen B., Gilchrist, E., Moerman, D. & Barstead, R. (2002). Improved detection of small deletions in complex pools of DNA. Nucleic Acids Research, 30 (12), e52.

McKay, S.J. & Jones, S.J.M. (2002). AcePrimer: automation of PCR primer design based on gene structure. Bioinformatics, 18 (11), 1538-1539.