Analysis of CYP81F2-A10, a potential candidate gene for the broad spectrum defense to Leptosphaeria maculans in Brassica napus L.
Canola (Brassica napus L.) is an economically important crop for producers in Canada and the ability to reduce disease causing yield loss is necessary for the longevity of the crop. A major disease that impacts canola production is blackleg, caused by the fungal pathogen Leptosphaeria maculans (Desm.) Ces. Et de Not. The understanding of how both quantitative and qualitative resistance to this disease works is vital to disease management. In this thesis, the response of the candidate gene CYP81F2-A10 (homolog of the Arabidopsis thaliana (L.) Heynh gene CYP81F2), and the 4-methoxy-indole-3yl-methyl-glucosinolate concentration upon infection examined using three B. napus near-isogenic lines (NILs) of Westar containing the allele CYP81F2-A10_Surpass400, and wild type, Westar containing the allele CYP81F2-A10_Westar. Plants were inoculated with either L. maculans (Isolate 87-41) or mock control (H2O). Relative to Westar, the NILs showed the first sign of suppression in pathogen growth and development at 6 days post inoculation (DPI). Full cDNA sequencing of CYP81F2-A10 revealed that CYP81F2-A10_Surpass400 contained a singular point mutation, resulting in an amino acid change. Analyses of transcript levels by RT-qPCR revealed that all lines showed upregulation of CYP81F2-A10 upon inoculation with L. maculans. The NILs showed gradual upregulation of the 4-methoxy-indole-3-yl-methyl-glucosinolate in response to inoculation with L. maculans, and two NILs (HN7 and NJ11) showed a significant difference from both their mock control treatment and inoculated Westar at 6 DPI. Westar did not show the same gradual increase in 4-methoxy-indole-3-yl-methyl-glucosinolate until 8DPI, where this increase did not correspond with pathogen suppression. This study suggests that the candidate gene CYP81F2-A10 is involved in the upregulation in 4-methoxy-indole-3-yl-methyl-glucosinolate in response to L. maculans inoculation in the early stages of infection. Furthermore, this type of interaction is associated with broad spectrum resistance and is non-specific to Avr genes.
Blackleg, L. Maculans, Brassica napus, Broad spectrum defense