CAN-TILL Projects

  For canola TILLING requests please go here:  .

In the future, we also hope to use this technique for reverse genetics in other crop species such as Camelina sativa, flax and safflower. We also envision using Ecotilling to identify natural genetic variation in species where this might be useful.

Past projects include Ecotilling in Populus trichocarpa (black cottonwood), and TILLING in Arabidopsis, the vegetable crop Brassica oleracea, and in the soil nematode Caenorhabditis elegans.

Current Projects	Past Projects
Brassica napus	Arabidopsis
	Brassica oleracea
	Caenorhabditis elegans
	Populus trichocarpa (Ecotilling)

Brassica napus TILLING

The Brassica project that we are currently working on is part of two oilseed initiatives funded by Genome Canada/Genome Alberta (Designing Oilseeds for Tomorrow's Markets), and AVAC (Bioactive Oils Programme). The goals of these projects are to use TILLING to identify endogenous mutations that affect oil content, seed coat characteristics and levels of anti-nutritional factors. It is anticipated that the results of this research will enhance the overall usefulness of canola seed, leading to improved meal for food and animal feed applications, and diversified seed oil content for nutritional and industrial uses. We are generating a population of approximately 6000 EMS-mutagenised B. napus lines for TILLING, and have identified mutations in a number of genes requested by DOTM and BOP researchers. The mutation rate for our population is greater than 1 mutation per 100Kb, a rate much higher than seen in Arabidopsis or other diploid species where TILLING has been used. This should allow the identification of several null or deleterious alleles for each gene screened and we intend to make this resource available to the general public upon completion of this project.

For canola TILLING requests, go here:  

Brassica oleracea TILLING

A past project, TILLING for mutations in the vegetable crop species Brassica oleracea, identified mutations that are thought to play a role in plant response to abiotic stresses that particularly affect crop yield in Canada. These include elements such as temperature, drought and nutrient imbalance. Phase one of this project involved TILLING in Arabidopsis homologues of Brassica genes that were thought to be important in abiotic stress responses in these crop species. The second phase of the project involved TILLING in Brassica oleracea, a species closely related to the commercially important crop, Brassica napus.

Photo from Brassica genetics for the classroom

C. elegans TILLING

Zetka MC, et al. 1999. Genes and Development 13:2258-2270.

C. elegans is a well-established model system ideal for genetic and molecular investigations into biological processes. The complete nematode genome sequence has been available for some time, and powerful in silico techniques have been developed for the prediction of gene function, expression and interaction. Despite the exciting possibilities flowing from predictions of structure/function relationships and the mapping of gene networks, the testing of these predictions relies largely on the existence of efficient reverse genetic approaches that target specific genes or classes of genes in vivo.

Our C. elegans pilot project is a collaboration with Monique Zetka, at McGill University, who has identified a target group of ~100 genes whose RNAi phenotype, expression profile, protein interaction data, or sequence homology has implicated them in genome surveillance, or chromosome segregation.

With assistance from Ann Rose and Nigel O'Neil at UBC, we have generated a library of purified genomic DNA from an EMS-mutagenised population of C. elegans and have demonstrated that the DNA can be used to effectively TILL for mutations in defined genes. We hope that the TILLING population we have generated will allow us to bring this valuable reverse genetic tool to the C. elegans research community in the near future.

Link to publications arising from this project:  

Ecotilling in poplar

The first tree to have it's genome sequenced is the western black cottonwood, Populus trichocarpa. This tree has a natural range that spans from Alaska to southern California, and from the Pacific Coast into interior mountain ranges in British Columbia, Washington and Oregon. As a first step towards analyzing genetic variation in this species, a live reference collection of P. trichocarpa has been established at the University of British Columbia (UBC) that includes trees from more than 140 different populations (see map).

We have used Ecotilling for the first time as a SNP discovery tool in a species that is long-lived, dioecious and genetically heterogeneous. SNP variation was examined at nine different loci in individuals from 41 different populations distributed throughout most of the P. trichocarpa range. Variation was analyzed both within a single tree (heterozygosity) as well as between individual trees and a reference, P. trichocarpa 383-2499 (Nisqually-1), whose genome has been sequenced. The availability of a sequenced genome made it possible to direct our attention to candidate genes of interest, providing an unprecedented view of genetic variation at multiple loci in this species. This pilot study shows that the level of nucleotide diversity in P. trichocarpa makes it theoretically possible to examine regions of 1000 to 1500 base pairs (bp) in 96 individuals or more on a single gel.

Link to publications arising from this project:  

This project was a collaboration with Quentin Cronk (Centre for Plant Research and Botanical Garden, UBC) and the Genome BC Forestry Genomics project.

Arabidopsis TILLING

Through collaboration with the Seattle TILLING Project, we used the model plant Arabidopsis thaliana as a tool for examining plant gene function through reverse genetics. We were able to provide researchers in the Genome Prairie Abiotic Stress group and the Genome B.C. Forestry Genomics group with a total of 171 mutations in 20 different genes that may be commercially important in agriculture or forestry. This project is completed.