Skip to main content
SpringerLink
Log in

Efficient marker-assisted breeding for clubroot resistance in elite Pol-CMS rapeseed varieties by updating the PbBa8.1 locus

  • Published:
Molecular Breeding Aims and scope Submit manuscript

Abstract

Clubroot disease poses a severe threat to rapeseed (Brassica napus) production worldwide and has recently been spreading across China at an unprecedented pace. Breeding and cultivation of resistant varieties constitute a promising and environment-friendly approach to mitigating this threat. In this study, the clubroot resistance locus PbBa8.1 was successfully transferred into SC4, a shared paternal line of three elite varieties in five generations by marker-assisted backcross breeding. Kompetitive allele specific PCR (KASP) markers of clubroot resistance gene PbBa8.1 and its linked high erucic acid gene (FAE1) were designed and applied for foreground selection, and 1,000 single-nucleotide polymorphisms (SNPs) were selected and used for the background selection. This breeding strategy produced recombinants with the highest recovery ratio of the recurrent parent genome (> 95%) at BC2F2 while breaking the linkage with FAE1 during the selection. An updated version of the paternal line (SC4R) was generated at BC2F3, showing significantly improved clubroot resistance at the seedling stage via artificial inoculation, and was comparable to that of the donor parent. Field trials of the three elite varieties and their updated versions in five environments indicated similar agronomic appearance and final yield. The introduced breeding strategy precisely pyramids the PbBa8.1 and FAE1 loci with the assistance of technical markers in a shorter period and could be applied to other desirable traits for directional improvement in the future.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

Data availability

Additional datasets generated by this study are included in the Supplementary Materials.

References

  • Bhattacharya I, Dutta S, Mondal S, Mondal B (2014) Clubroot disease on Brassica crops in India. Can J Plant Pathol 36(1):154–160

    Article  Google Scholar 

  • Chai AL, Xie XW, Shi YX, Li BJ (2014) Research status of clubroot (Plasmodiophora brassicae) on cruciferous crops in China. Can J Plant Pathol 36:142–153

    Article  Google Scholar 

  • Chen J, Jing J, Zhan Z, Zhang T, Zhang C, Piao Z et al (2013) Identification of novel QTLs for isolate-specific partial resistance to Plasmodiophora brassicae in Brassica rapa. PLoS One 8(12):e85307

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Chen ZJ, Tang DG, Ni JX, Li P, Wang L, Zhou JH et al (2021) Development of genic KASP SNP markers from RNA-Seq data for map-based cloning and marker-assisted selection in maize. BMC Plant Biol 21:157–167

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Cheon KS, Jeong YM, Oh H, Oh J, Kang DY, Kim N et al (2020) Development of 454 new kompetitive allele-specific PCR (KASP) markers for temperate japonica rice varieties. Plants 9:1531–1544

    Article  CAS  PubMed Central  Google Scholar 

  • Donald EC, Porter IJ (2014) Clubroot in Australia: the history and impact of Plasmodiophora brassicae in Brassica crops and research efforts directed towards its control. Can J Plant Pathol 36(1):66–84

    Article  CAS  Google Scholar 

  • Donald EC, Porter IJ, Lancaster RA (2001) Band incorporation of fluazinam (Shirlan) into soil to control clubroot of vegetable brassica crops. Aust J Exp Agric 41:1223–1226

    Article  CAS  Google Scholar 

  • Feng XM, Lin KX, Zhang WQ, Nan JZ, Zhang XH, Wang C et al (2019) Improving the blast resistance of the elite rice variety Kongyu-131 by updating the pi21 locus. BMC Plant Biol 19:249–260

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Han J, Lühs W, Sonntag K, Zähringer U, Borchardt DS, Wolter FP et al (2001) Functional characterization of β-ketoacyl-CoA synthase genes from Brassica napus L. Plant Mol Biol 46:229–239

    Article  CAS  PubMed  Google Scholar 

  • Hatakeyama K, Niwa T, Kato T, Ohara T, Kakizaki T, Matsumoto S (2017) The tandem repeated organization of NB-LRR genes in the clubroot resistant CRb, locus in Brassica rapa L. Mol Genet Genomics 292:397–405

    Article  CAS  PubMed  Google Scholar 

  • Hirai M, Harada T, Kubo N, Tsukada M, Suwabe K, Matsumoto S (2004) A novel locus for clubroot resistance in Brassica rapa and its linkage markers. Theor Appl Genet 108(4):639–643

    Article  CAS  PubMed  Google Scholar 

  • Hirani AH, Gao F, Liu J, Fu GH, Wu CR, McVetty PBE et al (2018) Combinations of independent dominant loci conferring clubroot resistance in all four turnip accessions (Brassica rapa) from the European clubroot differential set. Front Plant Sci 9:1628

    Article  PubMed  PubMed Central  Google Scholar 

  • Huang Z, Peng G, Gossen BD, Yu F (2019) Fine mapping of a clubroot resistance gene from turnip using SNP markers identified from bulked segregant RNA-Seq. Mol Breed 39:131–140

    Article  CAS  Google Scholar 

  • Karim MM, Dakouri A, Zhang Y, Chen Q, Yu F (2020) Two clubroot resistance genes, Rcr3 and Rcr9wa, mapped in Brassica rapa using bulk segregant RNA sequencing. Inter J Mol Sci 21(14):5033

    Article  CAS  Google Scholar 

  • Katsunori H, Keita S, Norio TR, Takeyuki K, Tsukasa N, Hiroyuki F et al (2013) Identification and characterization of Crr1a, a gene for resistance to clubroot disease Plasmodiophora brassicae Woronin) in Brassica rapa L. PLoS One 8:e54745

    Article  CAS  Google Scholar 

  • Kaur B, Mavi GS, Gill MS, Saini DK (2020) Utilization of KASP technology for wheat improvement. Cereal Res Commu 8057

  • Lander ES, Botstein D (1989) Mapping Mendelian factors underlying quantitative traits using RFLP linkage maps. Genetics 121(1):185–199

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Liu Z, Liu PW, Hong DF, He QB, Yang GS (2012) Fine mapping and candidate gene analysis of the nuclear restorer gene Rfp for pol CMS in rapeseed (Brassica napus). Theor Appl Genet 125(4):773–779

    Article  CAS  PubMed  Google Scholar 

  • Liu J, Qu J, Yang C, Tang D, Li J, Lan H et al (2015) Development of genome-wide insertion and deletion markers for maize, based on next-generation sequencing data. BMC Genomics 16(1):601

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Liu Z, Yang ZH, Wang X, Li KD, An H, Liu J et al (2016) A mitochondria-targeted PPR protein restores pol cytoplasmic male sterility by reducing orf224 transcript levels in oilseed rape. Mol Plant 9(7):1082–1084

    Article  CAS  PubMed  Google Scholar 

  • Lynch M, Milligan BG (1994) Analysis of population genetic structure with RAPD markers. Mol Ecol 3(2):91–99

    Article  CAS  PubMed  Google Scholar 

  • Malosetti M, Zwep LB, Forrest K, Eeuwijk FAV, Dieters M (2021) Lessons from a GWAS study of a wheat pre-breeding program: pyramiding resistance alleles to Fusarium crown rot. Theor Appl Genet 134(6):1–12

    Google Scholar 

  • Matsumoto E, Yasui C, Ohi M, Tsukada M (1998) Linkage analysis of RFLP markers for clubroot resistance and pigmentation in Chinese cabbage (Brassica rapa ssp. Pekinesis). Euphytica 104(2):79–86

    Article  CAS  Google Scholar 

  • Niwa R, Kumei T, Nomura Y, Yoshida S, Osaki M, Ezawa T (2007) Increase in soil PH due to Ca-rich organic matter application causes suppression of the clubroot disease of crucifers. Soil Biol Biochem 39:778–785

    Article  CAS  Google Scholar 

  • Pabinger S, Dander A, Fischer M, Snajder R, Sperk M, Efremova M et al (2014) A survey of tools for variant analysis of next-generation genome sequencing data. Brief Bioinform 15(2):256–278

    Article  PubMed  Google Scholar 

  • Pang WX, Liang S, Li XN, Li PP, Yu S, Lim YP et al (2014) Genetic detection of clubroot resistance loci in a new population of Brassica rapa. Hort Environ Biotechnol 55(6):540–547

    Article  CAS  Google Scholar 

  • Pang W, Fu P, Li X, Zhan Z, Yu S, Piao Z (2018) Identification and mapping of the clubroot resistance gene CRd in Chinese cabbage (Brassica rapa ssp. pekinensis). Front Plant Sci 9:653.

  • Patterson EI, Fleming MB, Kessler KC, Nissen SJ, Gaines TA (2017) A KASP genotyping method to identify northern watermilfoil, Eurasian watermilfoil, and their interspecific hybrids. Front Plant Sci 8:752

    Article  PubMed  PubMed Central  Google Scholar 

  • Piao ZY, Deng YQ, Choi SR, Park YJ, Lim YP (2004) SCAR and CAPS mapping of CRb, a gene conferring resistance to Plasmodiophora brassicae in Chinese cabbage (Brassica rapa ssp. Pekinesis). Theor Appl Genet 108(8):1458–1465

    Article  CAS  PubMed  Google Scholar 

  • Qu J, Liu J (2013) A genome-wide analysis of simple sequence repeats in maize and the development of polymorphism markers from next-generation sequence data. BMC Res Notes 6(1):403

    Article  PubMed  PubMed Central  Google Scholar 

  • Rafalski A (2002) Applications of single nucleotide polymorphisms in crop genetics. Curr Opin Plant Biol 5(2):94–100

    Article  CAS  PubMed  Google Scholar 

  • Rahman H, Peng G, Yu F, Falk KC, Kulkarni M, Selvaraj G (2014) Genetics and breeding for clubroot resistance in Canadian spring canola (Brassica napus L.). Can J Plant Pathol 36(1):122–134

    Article  CAS  Google Scholar 

  • Rasheed A, Wen W, Gao FM, Zhai SN, Jin H, Liu JD et al (2016) Development and validation of KASP assays for genes underpinning key economic traits in bread wheat. Theor Appl Genet 129(10):1843–1860

    Article  CAS  PubMed  Google Scholar 

  • Sakamoto K, Saito A, Hayashida N, Taguchi G, Matsumoto E (2008) Mapping of isolate-specific QTL for clubroot resistance in Chinese cabbage Brassica rapa L. ssp. pekinensis. Theo Appl Genet 117:759–767

    Article  CAS  Google Scholar 

  • Shah N, Sun J, Yu S, Yang Z, Wang Z, Huang F et al (2019) Genetic variation analysis of field isolates of clubroot and their responses to Brassica napus lines containing resistant gene CRb, PbBa8.1 and their combination inhomozygous and heterozygous state. Mol Breed 39:153.

  • Song YX, Duan XY, Wang PF, Li X, Yuan X, Wang ZY et al (2021) Comprehensive speed breeding: a high-throughput and rapid generation system for long-day crops. Plant Biotech J 10:1111

    Google Scholar 

  • Steele KA, Quinton-Tulloch MJ, Amgai RB, Dhakal R, Khatiwada SP, Vyas D et al (2018) Accelerating public sector rice breeding with high-density KASP markers derived from whole genome sequencing of Indica rice. Mol Breed 38:38–51

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Sun X, Pang H, Li M, Peng B, Guo H, Yan Q et al (2013) Evolutionary pattern of the FAE1 gene in Brassicaceae and its correlation with the erucic acid trait. PLoS One 8:e83535

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Suwabe K (2006) Simple sequence repeat-based comparative genomics between Brassica rapa and Arabidopsis thaliana: the genetic origin of clubroot resistance. Genetics 173(1):309–319

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Suwabe K, Tsukazaki H, Iketani H, Hatakeyama K, Fujimura M, Nunome T et al (2003) Identification of two loci for resistance to clubroot (Plasmodiophora brassicae Woronin) in Brassica rapa L. Theor Appl Genet 107(6):997–1002

    Article  CAS  PubMed  Google Scholar 

  • Swathi G, Rani CVD, Md J, Madhav MS, Vanisree S, Anuradha Ch et al (2019) Marker-assisted introgression of the major bacterial blight resistance genes, Xa21 and xa13, and blast resistance gene, Pi54, into the popular rice variety, JGL1798. Mol Breed 39:58

    Article  CAS  Google Scholar 

  • Tian E, Liang H, Wang J, Guo J, Cao H, Lin S (2018) Variation and correlation of erucic acid, oleic acid and glucosinolate contents in Brassica rapa seeds. Asian Agric Res 10:57–60

    Google Scholar 

  • Vos P, Hogers R, Bleeker M, Reijans M, Tvd L, Hornes M et al (1995) AFLP: a new technique for DNA fingerprinting. Nucleic Acids Res 23(21):4407–4414

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Wallenhammar AC, Almquist C, Schwelm A, Roos J, Marzec-Schmidt K, Jonsson A, et al (2014) Clubroot, a persistent threat to Swedish oilseed rape production. Can J Plant Pathol 135–141

  • Wang TH, Li B, Guo YM, Liu XH, Li B, Peng P et al (2020a) Technical regulation for Pol CMS hybrids purity identification of Brassica napus L. by KASP method. HNZ267–2020a, Department of agriculture and rural areas, Hunan Provinces.

  • Wang D, Yang T, Liu R, Li NN, Wang XW, Ashutosh S et al (2020b) RNA-Seq analysis and development of SSR and KASP markers in lentil (Lens culinaris Medikus subsp. culinaris). Crop J 8:953–965

    Article  Google Scholar 

  • Wang TH, Li B, Guo YM, Zhang YT, Liu XH, Fan LY et al (2020c) Development of SSR markers associated with Polima CMS and its application. Mol Plant Breed 11:10

    Google Scholar 

  • Wang YY, Yang ZQ, Yang QY, Zhang CY (2021) Genetic improvement and application of clubroot resistance in Brassica napus. J Huazhong Agric Uni 40(2):1–5

    CAS  Google Scholar 

  • Williams PH (1966) A system for the determination of races of Plasmodiophora brassicae that infect cabbage and rutabaga. Phytopathol 56:624–626

    Google Scholar 

  • Xiao Q, Wang HD, Song N, Yu Z, Khan I, Xie WB et al (2021) The Bnapus50K array: a quick and versatile genotyping tool for Brassica napus genomic breeding and research. G3 11(10):jkab241

    Article  PubMed  PubMed Central  Google Scholar 

  • Yang GL, Chen SP, Chen LK, Sun K, Huang CH, Zhou DH et al (2019) Development of a core SNP arrays based on the KASP method for molecular breeding of rice. Rice 12(1):21

    Article  PubMed  PubMed Central  Google Scholar 

  • Zhan ZX, Nwafor CC, Hou ZK, Gong JF, Zhu BB, Jiang YF et al (2017) Cytological and morphological analysis of hybrids between Brassicoraphanus and Brassica napus for introgression of clubroot resistant trait into Brassica napus L. PLoS One 12:e0177470

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Zhan ZX, Jiang YF, Shah N, Hou ZK, Zhou YW, Dun BC et al (2020) Association of clubroot resistance locus PbBa8.1 with a linkage drag of high erucic acid content in the seed of the European turnip. Front Plant Sci 11:810

    Article  PubMed  PubMed Central  Google Scholar 

  • Zhang T, Zhao Z, Zhang CY, Pang WX, Choi SR, Lim YP, Piao ZY (2014) Fine genetic and physical mapping of the CRb gene conferring resistance to clubroot disease in Brassica rapa. Mol Breed 34:1173–1183

    Article  CAS  Google Scholar 

Download references

Acknowledgements

The clubroot-resistant donor parent was generously shared by Prof. Chunyu Zhang. We appreciate Prof. Zhongyun Piao for earlier examination of pathotypes from the Hunan clubroot-infected region. We are deeply thankful for the assistance with agronomic evaluation and field sampling provided by the five local agricultural research institutes of Changsha, Changde, Hengyang, Jishou, and Huaihua.

Funding

This project was supported by grants from the National Key Research and Development Program of China (Grant No. 2021YFD1600500), Natural Science Foundation for Young Scientists of Hunan Province (Grant No. 2021JJ40330), Changsha Municipal Natural Science Foundation (Grant No. kq2014179), and the China Agricultural Research System (CARS-12).

Author information

Author notes

  1. Yiming Guo and Bao Li contributed equally to this work.

Authors and Affiliations

  1. Crop Research Institute, Hunan Academy of Agricultural Sciences, Changsha, 410125, China

    Yiming Guo, Bao Li, Mei Li, Qian Yang, Xinhong Liu, Liang Qu, Lianyi Fan & Tonghua Wang

  2. Hunan Engineering and Technology Research Center of Hybrid Rapeseed, Changsha, 410125, China

    Yiming Guo, Bao Li, Mei Li, Qian Yang, Xinhong Liu, Liang Qu, Lianyi Fan & Tonghua Wang

  3. Hunan Provincial Key Laboratory for Biology and Control of Plant Diseases and Insect Pests, Hunan Agricultural University, Changsha, 410128, China

    Hongjian Zhu

Authors
  1. Yiming Guo
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Bao Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Mei Li
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Hongjian Zhu
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Qian Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Xinhong Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Liang Qu
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Lianyi Fan
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Tonghua Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

Contributions

Yiming Guo and Bao Li conducted the experiments and drafted the manuscript. Mei Li, Liang Qu, and Lianyi Fan were involved in the agronomic evaluation of the previous and updated varieties. Qian Yang took part in data analysis and plotting. Hongjian Zhu and Xinhong Liu participated in the artificial inoculation of seedlings. Tonghua Wang designed the experiment and had oversight of the entire process. All authors read and approved the final manuscript.

Corresponding author

Correspondence to Tonghua Wang.

Ethics declarations

Ethics approval and consent to participate

Not applicable.

Consent for publication

The authors consented for the publication.

Competing interests

The authors declare no competing interests.

Additional information

Publisher's note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Below is the link to the electronic supplementary material.

Supplementary file1 (XLSX 14 KB)

Supplementary file2 (DOCX 95 KB)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Guo, Y., Li, B., Li, M. et al. Efficient marker-assisted breeding for clubroot resistance in elite Pol-CMS rapeseed varieties by updating the PbBa8.1 locus. Mol Breeding 42, 41 (2022). https://doi.org/10.1007/s11032-022-01305-9

Download citation

  • Received:

  • Accepted:

  • Published:

  • DOI: https://doi.org/10.1007/s11032-022-01305-9

Keywords

Search

Navigation