The protist Plasmodiophora brassicae is a biotrophic soil-borne pathogen that causes clubroot on Brassica crops worldwide. Clubroot disease is a serious threat to the 8 M ha of canola (Brassica napus) grown annually in western Canada. While host resistance is the key to clubroot management, sources of resistance are limited.

Clubroot, caused by Plasmodiophora brassicae, is an important disease of Brassica crops worldwide. F₁ progeny from the Brassica rapa lines T19 (resistant) × ACDC (susceptible) were backcrossed with ACDC, then self-pollinated to produce BC₁S₁ lines, From genotyping-by-sequencing (GBS) of the parental lines and BC₁ plants, about 1.32 M sequences from T19 were aligned into the reference genome of B. rapa with 0.4-fold coverage, and 1.77 M sequences with 0.5-fold coverage in ACDC. The number of aligned short reads per plant in the BC₁ ranged from 0.07 to 1.41 M sequences with 0.1-fold coverage. A total of 1584 high quality SNP loci were obtained, distributed on 10 chromosomes. A single co-localized QTL, designated as Rcr4 on chromosome A03, conferred resistance to pathotypes 2, 3, 5, 6 and 8. The peak was at SNP locus A03_23710236, where LOD values were 30.3 to 38.8, with phenotypic variation explained (PVE) of 85–95%. Two QTLs for resistance to a novel P. brassicae pathotype 5x, designated Rcr8 on chromosome A02 and Rcr9 on A08, were detected with 15.0 LOD and 15.8 LOD, and PVE of 36% and 39%, respectively. Bulked segregant analysis was performed to examine TIR-NBS-LRR proteins in the regions harboring the QTL.

Clubroot, caused by Plasmodiophora brassicae, is an important disease of canola (Brassica napus) in western Canada and worldwide. In this study, a clubroot resistance gene (Rcr2) was identified and fine mapped in Chinese cabbage cv. ‘Jazz’ using SNP markers identified from bulked segregant RNA sequencing (BSR-Seq) and molecular markers were developed for use in marker assisted selection. In total, 203.9 million raw reads were generated from one pooled resistant (R) and one pooled susceptible (S) sample, and >173,000 polymorphic single nucleotide polymorphisms (SNP) sites were identified between the R and S samples. One significant peak was observed between 22-26 Mb of chromosome A03, which had been predicted by BSR-Seq to contain the causal gene Rcr2. There were 490 polymorphic SNP sites identified in the region. A segregating population consisting of 675 plants was analysed with 15 SNP sites in the region using the Kompetitive Allele Specific PCR method, and Rcr2 was fine mapped between two SNP markers, SNP_A03_32 and SNP_ A03_67 with 0.1 and 0.3 centi-Morgan from Rcr2, respectively. Five SNP markers co-segregated with Rcr2 in this region. Variants were identified in 14 of 36 genes annotated in the Rcr2 target region. The numbers of poly variants differed among the genes. Four genes encode TIR-NBS-LRR proteins and two of them Bra019410 and Bra019413, had high numbers of polymorphic variants and so are the most likely candidates of Rcr2.

Genetic resistance is widely used to manage clubroot (Plasmodiophora brassicae) in brassica crops, but new pathotypes have recently been identified on canola (Brassica napus) on the Canadian prairies. Resistance effective against both the most prevalent pathotype (3H, based on the Canadian Clubroot Differential system) and the new pathotypes is needed. BC₁ plants of Brassica rapa from a cross of line 96-6990-2 (clubroot resistance originating from turnip cultivar ‘Waaslander’) and a susceptible doubled-haploid line, ACDC, exhibited a 1:1 segregation for resistance against pathotypes 3H and 5X. A resistance gene designated as Rcr3 was mapped initially based on the percentage of polymorphic variants using bulked segregant RNA sequencing (BSR-Seq) and further mapped using Kompetitive Allele Specific PCR. DNA variants were identified by assembling short reads against a reference genome of B. rapa. Rcr3 was mapped into chromosome A08. It was flanked by single nucleotide polymorphisms (SNP) markers (A90_A08_SNP_M12 and M16) between 10.00 and 10.23 Mb, in an interval of 231.6 Kb. There were 32 genes in the Rcr3 interval. Three genes (Bra020951, Bra020974, and Bra020979) were annotated with disease resistance mechanisms, which are potential candidates for Rcr3. Another resistance gene, designated as Rcr9^wa, for resistance to pathotype 5X was mapped, with the flanking markers (A90_A08_SNP_M28 and M79) between 10.85 and 11.17 Mb using the SNP sites identified through BSR-Seq for Rcr3. There were 44 genes in the Rcr9^wa interval, three of which (Bra020827, Bra020828, Bra020814) were annotated as immune-system-process related genes, which are potential candidates for Rcr9^wa.

Clubroot, caused by Plasmodiophora brassicae, is an important disease on Brassica species worldwide. A clubroot resistance gene, Rcr1, with efficacy against pathotype 3 of P. brassicae, was previously mapped to chromosome A03 of B. rapa in pak choy cultivar “Flower Nabana”. In the current study, resistance to pathotypes 2, 5 and 6 was shown to be associated with Rcr1 region on chromosome A03. Bulked segregant RNA sequencing was performed and short read sequences were assembled into 10 chromosomes of the B. rapa reference genome v1.5. For the resistant (R) bulks, a total of 351.8 million (M) sequences, 30,836.5 million bases (Mb) in length, produced 120-fold coverage of the reference genome. For the susceptible (S) bulks, 322.9 M sequences, 28,216.6 Mb in length, produced 109-fold coverage. In total, 776.2 K single nucleotide polymorphisms (SNPs) and 122.2 K insertion / deletion (InDels) in R bulks and 762.8 K SNPs and 118.7 K InDels in S bulks were identified; each chromosome had about 87% SNPs and 13% InDels, with 78% monomorphic and 22% polymorphic variants between the R and S bulks. Polymorphic variants on each chromosome were usually below 23%, but made up 34% of the variants on chromosome A03. There were 35 genes annotated in the Rcr1 target region and variants were identified in 21 genes. The numbers of poly variants differed significantly among the genes. Four out of them encode Toll-Interleukin-1 receptor / nucleotide-binding site / leucine-rich-repeat proteins; Bra019409 and Bra019410 harbored the higher numbers of polymorphic variants, which indicates that they are more likely candidates of Rcr1. Fourteen SNP markers in the target region were genotyped using the Kompetitive Allele Specific PCR method and were confirmed to associate with Rcr1. Selected SNP markers were analyzed with 26 recombinants obtained from a segregating population consisting of 1587 plants, indicating that they were completely linked to Rcr1. Nine SNP markers were used for marker-assisted introgression of Rcr1 into B. napus canola from B. rapa, with 100% accuracy in this study.

Resistance to the blackleg disease of Brassica napus (canola/oilseed rape), caused by the hemibiotrophic fungal pathogen Leptosphaeria maculans, is determined by both race-specific resistance (R) genes and quantitative resistance loci (QTL), or adult-plant resistance (APR). While the introgression of R genes into breeding material is relatively simple, QTL are often detected sporadically, making them harder to capture in breeding programs. For the effective deployment of APR in crop varieties, resistance QTL need to have a reliable influence on phenotype in multiple environments and be well defined genetically to enable marker-assisted selection (MAS). Doubled-haploid populations produced from the susceptible B. napus variety Topas and APR varieties AG-Castle and AV-Sapphire were analysed for resistance to blackleg in two locations over 3 and 4 years, respectively. Three stable QTL were detected in each population, with two loci appearing to be common to both APR varieties. Physical delineation of three QTL regions was sufficient to identify candidate defense-related genes, including a cluster of cysteine-rich receptor-like kinases contained within a 49 gene QTL interval on chromosome A01. Individual L. maculans isolates were used to define the physical intervals for the race-specific R genes Rlm3 and Rlm4 and to identify QTL common to both field studies and the cotyledon resistance response. Through multi-environment QTL analysis we have identified and delineated four significant and stable QTL suitable for MAS of quantitative blackleg resistance in B. napus, and identified candidate genes which potentially play a role in quantitative defense responses to L. maculans.