title: Identification of Genome-Wide Variants and Discovery of Variants Associated with Brassica rapa Clubroot Resistance Gene Rcr1 through Bulked Segregant RNA Sequencing.
abstract: 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.
authors.affiliation: Saskatoon Research Centre, Agriculture and Agri-Food Canada, Saskatoon, Saskatchewan, Canada.
authors.forename: Fengqun
authors.initials: F
authors.lastname: Yu
authors.affiliation: Saskatoon Research Centre, Agriculture and Agri-Food Canada, Saskatoon, Saskatchewan, Canada.
authors.forename: Xingguo
authors.initials: X
authors.lastname: Zhang
authors.affiliation: Saskatoon Research Centre, Agriculture and Agri-Food Canada, Saskatoon, Saskatchewan, Canada.
authors.forename: Zhen
authors.initials: Z
authors.lastname: Huang
authors.affiliation: Saskatoon Research Centre, Agriculture and Agri-Food Canada, Saskatoon, Saskatchewan, Canada.
authors.forename: Mingguang
authors.initials: M
authors.lastname: Chu
authors.affiliation: Saskatoon Research Centre, Agriculture and Agri-Food Canada, Saskatoon, Saskatchewan, Canada.
authors.forename: Tao
authors.initials: T
authors.lastname: Song
authors.affiliation: Saskatoon Research Centre, Agriculture and Agri-Food Canada, Saskatoon, Saskatchewan, Canada.
authors.forename: Kevin C
authors.initials: KC
authors.lastname: Falk
authors.affiliation: Department of Plant Agriculture, University of Guelph, Guelph, Ontario, Canada.
authors.forename: Abhinandan
authors.initials: A
authors.lastname: Deora
authors.affiliation: Saskatoon Research Centre, Agriculture and Agri-Food Canada, Saskatoon, Saskatchewan, Canada.
authors.forename: Qilin
authors.initials: Q
authors.lastname: Chen
authors.affiliation: Saskatoon Research Centre, Agriculture and Agri-Food Canada, Saskatoon, Saskatchewan, Canada.
authors.forename: Yan
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Saskatoon Research Centre, Agriculture and Agri-Food Canada, Saskatoon, Saskatchewan, Canada.
authors.forename: Linda
authors.initials: L
authors.lastname: McGregor
authors.affiliation: Saskatoon Research Centre, Agriculture and Agri-Food Canada, Saskatoon, Saskatchewan, Canada.
authors.forename: Bruce D
authors.initials: BD
authors.lastname: Gossen
authors.affiliation: Department of Plant Agriculture, University of Guelph, Guelph, Ontario, Canada.
authors.forename: Mary Ruth
authors.initials: MR
authors.lastname: McDonald
authors.affiliation: Saskatoon Research Centre, Agriculture and Agri-Food Canada, Saskatoon, Saskatchewan, Canada.
authors.forename: Gary
authors.initials: G
authors.lastname: Peng
mesh_terms: D001483:Base Sequence; D029741:Brassica rapa / Q000235:genetics* / Q000469:parasitology; D032461:Chromosomes, Plant / Q000235:genetics; D060467:Disease Resistance / Q000235:genetics*; D005787:Gene Frequency; D017343:Genes, Plant / Q000235:genetics*; D018745:Genome, Plant / Q000235:genetics*; D005838:Genotype; D054643:INDEL Mutation; D008969:Molecular Sequence Data; D010935:Plant Diseases / Q000235:genetics / Q000469:parasitology; D018517:Plant Roots / Q000235:genetics* / Q000469:parasitology; D056909:Plasmodiophorida / Q000502:physiology; D020641:Polymorphism, Single Nucleotide; D017423:Sequence Analysis, RNA / Q000379:methods