Exploiting susceptibility genes in canola to improve blackleg resistance

Term: 3 years, beginning in 2023
Status: Ongoing
Researcher(s): Gary Peng, AAFC
SaskCanola Investment: $81,125
Total Project Cost: $298,250
Funding Partners: ACPC, MCGA, WGRF

Objectives

The proposed research will explore a novel approach to identify susceptibility genes in canola and develop additional CRISPR/cas9 knock-out lines with unique blackleg resistance traits for canola breeding.

1) Assess blackleg resistance levels in the TILLING population available at GIFS and UBC-13 mutants available at AAFC Saskatoon;

2) Identify candidate S genes conferring blackleg resistance, and develop associated molecular markers;

3) Generate stable S gene knock-out canola lines using CRISPR/Cas9 technology, evaluate their resistance reaction, develop homozygous pre-breeding resistant lines for tech transfer.

Project Description

This proposed research aims to study new resistance modes of action based on S genes in canola and potentially develop novel R resource for sustainable management of blackleg in western Canada. This study will tap into a unique diversity-rich TILLING canola population (Gilchrist et al. 2013) and UBC-13 mutants available respectively at GIFS and AAFC Saskatoon, to discover S-related genes and their desirable variants for broad-spectrum/stable blackleg resistance. The assessment of these canola lines (modified/mutagenized) will help to identify and characterize candidate S genes that promote blackleg infection in canola. Then we will generate homozygous S genes knock-out canola lines using CRISPR/Cas9 technology to generate novel resources and markers for blackleg resistance to be used in breeding for canola varieties with potentially broad-spectrum and durable disease resistance traits. This approach in exploiting the TILLING, UBC-13 and other resources may also be useful for resistance against other canola diseases in western Canada. The potential benefit of this research to producers and industry is the continued improvement of variety resistance to all important canola diseases in western Canada.

Blackleg is the most widely occurred diseases on canola in western Canada, causing not only yield losses, but also trade disputes in export. Amongst all the measures recommended for integrated blackleg management, genetic resistance is a cornerstone, largely pivoting on identification and deployment of specific resistance (R) genes and quantitative resistance (QR) in breeding materials. However, there are only limited number of R genes available for blackleg resistance breeding, and for almost every R gene known, there is at least one virulent pathogen race existing in western Canada already. Therefore, it is only the matter of time for any R gene to be overcome once it is introduced. QR has been an important element for blackleg resistance in most of the Canadian cultivars, but it is very difficult to improve it due to its multi-genic nature.

Resistance to crop diseases can also be achieved via the loss-of-function of susceptibility (S) genes, i.e. through a functional disruption of S genes, because these S genes provide a base for compatibility with a pathogen during host-pathogen interactions, allowing or facilitating the infection process. This proposed study explores potential new S genes related to virulence of blackleg pathogen and severity of the disease. A two-step strategy will be used by starting with the screening of a unique canola TILLING (Targeting Induced Local Lesions in Genomes) population available at the Global Institute for Food Security (GIFS) to identify prospective S gene candidates. Additional canola mutants developed from the susceptible canola cultivar “Westar” with knocked-out(s) for genes encoding ubiquitin conjugating enzyme (UBC-13), a key pathogenicity factor, will also be evaluated. The studies performed with mutants generated using a CRISPR/Cas9-based platform at AAFC Saskatoon and have shown encouraging results displaying a range of resistance phenotypes to clubroot. Once promising candidates have been identified towards blackleg, canola mutants with targeted loss-of-function S genes will be generated using the CRISPR/Cas9 technology for development of more lines with broad-spectrum and stable resistance against blackleg. Success of this approach and resources developed may also be applicable and useful to other canola diseases, including clubroot and sclerotinia stem rot.