QTL mapping and transcriptome analysis identify novel QTLs and candidate genes in Brassica oleracea var. villosa for quantitative resistance against Sclerotinia sclerotiorum

doi:10.21203/rs.3.rs-2312444/v1

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QTL mapping and transcriptome analysis identify novel QTLs and candidate genes in Brassica oleracea var. villosa for quantitative resistance against Sclerotinia sclerotiorum

https://doi.org/10.21203/rs.3.rs-2312444/v1

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published 26 Mar, 2023

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Sclerotinia stem rot (SSR), caused by Sclerotinia sclerotiorum, is one of the most destructive diseases in oilseed rape growing regions. To date, there is no effective genetic resistance against S. sclerotiorum in the B. napus germplasm and knowledge of the molecular plant-fungal interaction is also limited. To identify new resistance resources, we screened a set of wild Brassica species and identified B. villosa (BRA1896) with a high Sclerotinia-resistance. Two segregating F2 populations for Sclerotinia-resistance, generated by interspecific crosses between the resistant B. villosa (BRA1896) and the wild susceptible B. oleracea (BRA1909) were assessed for Sclerotinia-resistance. Genetic mapping using a 15k Illumina Infinium SNP- array resulted in a high-density genetic map containing 1,118 SNP markers and spanning a total genetic length of 792.2 cM. QTL analysis revealed seven QTLs explaining 3.8 % to 16.5 % of phenotypic variance. Intriguingly, RNAseq-based transcriptome analysis identified genes and pathways specific to B. villosa, of which a cluster of five genes encoding putative receptor-like kinases (RLKs) and two pathogenesis-related (PR) proteins are co-localized within a QTL on chromosome C07. Furthermore, transcriptomic analysis revealed enhanced ethylene (ET)-activated signaling in the resistant B. villosa, which is associated with a stronger plant immune response, depressed cell death, and enhanced phytoalexin biosynthesis compared to the susceptible B. oleracea. Our data demonstrate that B. villosa represents a novel and unique genetic source for improving oilseed rape resistance against SSR.

Tables 1 to 3 are available in the Supplementary Files section.

TablesSclerotiniaBergmannetal.2022.pdf
Table 1 Identified QTLs for Sclerotinia-resistance in the wild Brassica populations. Table 2 De novo assembled genes specifically expressed in B. villosa (BRA1896) and associated with plant defense response. Table 3 Pathogenesis-related (PR) genes in the identified QTLs in the B. oleracea TO1000 reference genome.
SupplementaryFiguresBergmannetal.2022.pdf
Figure S1 Trypan blue staining of inoculated leaves of A – B B. oleracea (BRA1909) and C – D B. villosa (BRA1896). A dense and compact structural fungal growth mainly within necrotic tissue was observed in leaves of the susceptible B. oleracea. In the resistant B. villosa, fungal expansion appeared less structured and focused mainly on the leaf surface with no delimited junction between healthy and infected tissue. Figure S2 Venn diagram showing the proportion of F2 individuals in A Population A and B Population B which were commonly classified as resistant, intermediate, and susceptible in the leaf- and petiole-assay. The grouping is based on the comparison to the resistant B. villosa (BRA1896) and the susceptible B. oleracea (BRA1909) parents in each population, respectively. Figure S3 Lesion size distributions of genotyped F2 plants. A – D Leaf-lesion size distributions of all genotyped F2 individuals in A Population A and B – D in Population B. E – H Petiole-lesion size distributions of all genotyped F2individuals in E Population A and F – H in Population B. Figure S4 Logarithm of the odds (LOD) profiles in both mapping populations. Horizontal lines indicate the threshold for significance. Different assays are highlighted by different colors. Figure S5 Principal component analysis (PCA) of RNAseq samples of B. villosa (BRA1896) and B. oleracea (BRA1909) after regularized log-transformation of the count matrix in DESeq2. Biological replications are highlighted by identical colors. PC1: variance between samples caused by species, PC2: variance between samples caused by the treatment. Figure S6 Marker gene expression comparison of reverse-transcribed quantitative PCR (RT-qPCR) and RNAseq data of resistant (BRA1896) and susceptible (BRA1909) petioles at 8 hours post inoculation (hpi) with Sclerotinia. Asterisks indicate a significant induction compared to the control sample. BRA1896 = B. villosa, BRA1909 = B. oleracea. RT-qPCR expression was log2-transformed for a better comparison to the RNAseq. Genes in the RNAseq were as follows: BolETR2 = Unigene.2465, BolAOC3 = Bo9g075870, BolLOX3 = Bo8g067210, BolPDF1.2 = Bo2g086460, BolNCED3 = Bo5g130280, BolPR1 = Bo3g088360. Primers for RT-qPCR analysis are listed in Supplementary Table S1.
SupplementaryTablesBergmannetal.2022.pdf
Table S1 Primer used for real-time quantitative PCR gene expression analysis. Table S2 Summary of the genetic map. Table S3 Sequencing output and mapping rates for RNAseq samples of B. villosa (BRA1896) and B. oleracea (BRA1909). Table S4 Specifically expressed genes involved in the immune response of B. villosa. Table S5 Ethylene-associated genes (iDEGs) with a significant difference in their expression between 989 B. villosa (BRA1896) and B. oleracea (BRA1909).
SupplementaryDataBergmannetal.2022.xlsx

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Journal Publication

published 26 Mar, 2023

Read the published version in Theoretical and Applied Genetics →

Editorial decision: Major revisions
14 Feb, 2023
Reviewers agreed at journal
11 Dec, 2022
Reviewers invited by journal
11 Dec, 2022
Editor assigned by journal
06 Dec, 2022
First submitted to journal
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You are reading this latest preprint version

QTL mapping and transcriptome analysis identify novel QTLs and candidate genes in Brassica oleracea var. villosa for quantitative resistance against Sclerotinia sclerotiorum

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