Transcriptomic responses in spring canola carrying clubroot resistance introgressed from rutabaga or “Mendel”

• Author / Creator

  Summanwar, Aarohi

• Canola provides ~15% of the total global vegetable oil supply and is an important crop contributing about $26.7 billion to Canada’s economy. Canola production has been threatened by clubroot disease caused by Plasmodiophora brassicae Woronin, which in extreme case can result complete crop failure. Development of clubroot resistant cultivars is one of the important ways of managing this disease. Clubroot resistance has been introgressed into spring canola from various sources of which the resistance introgressed from winter canola cv. ‘Mendel’ and rutabaga have been used most extensively in Canada; however, the molecular basis including comparative analysis of this resistance has not yet been studied. In this dissertation, I focused and compared thetranscriptomic response of these two resistances in canola in response to infection by P. brassicae.
  Eukaryote transcriptome is more complex than previously imagined, as evidenced by the knowledge gained from recent studies. The transcriptomes primarily consist of non-coding RNAs such as long non-coding RNAs (lncRNAs) and protein coding RNAs like mRNAs. Using lncRNA-seq, we identified 530 differentially expressed (DE) lncRNAs between clubroot resistant canola lines carrying resistance of rutabaga (RtR) and susceptible lines lacking this resistance (RtS). Similarly, we also identified 464 DE lncRNAs between the canola lines carrying resistance of winter cv. ‘Mendel’ (MdR) and lines lacking this resistance (MdS). While comparing these two sets of DE lncRNAs, we found 12 lncRNAs were upregulated in both the resistances and could be potentially involved in regulating clubroot resistance. We also found that the predicted targets genes of these lncRNAs were similar in both RtR and MdR resistances. Quantitative real-time (qRT-PCR) analysis was carried out for the lncRNAs specifically expressed in RtR or MdR lines, and the lncRNAs expressed in both RtR and MdR lines and their respective target genes to confirm their response to clubroot .
  Pathway analysis of the target genes regulated by the 530 DE lncRNAs identified by using rutabaga-resistance and 464 DE lncRNAs identified by using Mendel-resistance showed that the primary and secondary metabolic pathways play an important role in resistance to this disease at 10 dpi; the genes involved in primary and secondary metabolic pathways were observed to be upregulated in the RtR and MdR lines. The importance of these pathways was investigated by treating, clubroot susceptible plants with trehalose. The DH lines treated with 30 mM and 60 mM trehalose showed partial resistance to this disease while the treatment with 120 mM trehalose did not change the susceptible phenotype. Additionally, we observed that the expression of the genes like pathogenesis-related protein 2, lipoxygenase and phenylalanine ammonia lyase increased in response to this treatment indicating that the modulation of primary metabolic pathways can affect clubroot resistance.
  Furthermore, considering the importance of the clubroot resistance loci of BnaA03 from MdR lines and BnaA08 from RtR lines and their association with clubroot resistance, the DEGs upregulated from these two chromosomes were compared, and the gene cytokinin responsive factor (CRF4) was found upregulated in both resistances and, thus, could be involved in mediating clubroot resistance in both RtR and MdR lines. In addition, I also developed lncRNA- and gene-based molecular markers from BnaA08 (RtR) and BnaA03 (MdR). A simple sequence repeat (SSR) marker designed from the lncRNA LNC_000424 showed cosegregation with resistance in a mapping population carrying Mendel-resistance; this lncRNA was upregulated in the MdR lines and was predicted to regulate the target genes involved in plant defense. Additionally, the gene-based SSR markers from the DEGs viz. BnaA03g41300D and BnaA03g44400D of BnaA03 upregulated in the MdR lines and SSR from the DEG BnaA08g03250D of BnaA08 upregulated in the RtR lines showed linkage association with resistance; however, these lncRNA- and gene-based markers showed 1-3% recombination between the marker and resistance phenotypes.

• Subjects / Keywords

  • lncRNA
  • Canola
  • Transcriptomics
  • lncRNA-seq
  • Clubroot

• Graduation date

  Fall 2020

• Type of Item

  Thesis

• Degree

  Doctor of Philosophy

• DOI

  https://doi.org/10.7939/r3-j6wa-0766

• License

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