RESUMO
The clubroot disease has become a worldwide threat for crucifer crop production, due to its soil-borne nature and difficulty to eradicate completely from contaminated field. In this study we used an elite resistant European fodder turnip ECD04 and investigated its resistance mechanism using transcriptome, sRNA-seq, degradome and gene editing. A total of 1751 DEGs were identified from three time points after infection, among which 7 hub genes including XTH23 for cell wall assembly and two CPK28 genes in PTI pathways. On microRNA, we identified 17 DEMs and predicted 15 miRNA-target pairs (DEM-DEG). We validated two pairs (miR395-APS4 and miR160-ARF) by degradome sequencing. We investigated the miR395-APS4 pair by CRISPR-Cas9 mediated gene editing, the result showed that knocking-out APS4 could lead to elevated clubroot resistance in B. napus. In summary, the data acquired on transcriptional response and microRNA as well as target genes provide future direction especially gene candidates for genetic improvement of clubroot resistance on Brassica species.
RESUMO
Clubroot is caused by Plasmodiophora brassicae, which threatens Brassicaceae crop production worldwide. In recent years, there has been an outbreak and rapid spread of clubroot in many major cruciferous crop-producing areas of China. In this study, we identified a cabbage material DingWen (DW) with different resistant capabilities from Huashuang5R (H5R) and Huayouza62R of Brassica napus, which are currently used as the main resistant cultivars for clubroot management in China. We used a next-generation sequencing-based bulked segregant analysis approach, combined with genetic mapping to identify clubroot-resistant (CR) genes from F1 population generated from a cross between the DW (CR) and HZSX (clubroot susceptible). The CR locus of DW (named CRA8.1) was mapped to a region between markers A08-4346 and A08-4853, which contains two different loci CRA8.1a and CRA8.1b after fine mapping. The CRA8.1b loci contain a fragment of 395 kb between markers A08-4624 and A08-4853 on A08 chromosome, and it is responsible for the resistance to PbZj and PbXm isolates. However, together with CRA8.1a, corresponding to a 765-kb region between markers A08-4346 and A08-4624, then it can confer resistance to PbXm +. Finally, through expression analysis between resistant and susceptible materials, two genes encoding TIR-NBS-LRR proteins (BraA08g039211E and BraA08g039212E) and one gene encoding an RLP protein (BraA08g039193E) were identified to be the most likely CR candidates for the peculiar resistance in DW.
