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Identification of Candidate Genes Associated With Resistance Against Race 0 of Colletotrichum lentis In Lens ervoides

Date

2020-02-24

Journal Title

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Publisher

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Type

Thesis

Degree Level

Masters

Abstract

Lens ervoides is a potential source of resistance to anthracnose caused by the pathogen Colletotrichum lentis. Transcriptome sequencing was performed on the resistant LR-66-528 and susceptible LR-66-524 recombinant inbred lines (RILs) of L. ervoides infected with the aggressive race 0 isolate CT-30 of C. lentis to unravel the genetic control underlying the genetic responses against this pathogen. The inoculated samples were harvested at 6, 12, 24, 48, 72, 96 and 144 hours post-inoculation (hpi) for molecular studies. Results of quantitative PCR to estimate fungal biomass revealed that 24, 48, 72 and 96 hpi were interesting time-points for studying disease development because of exponential trends of fungal growth during this period. Subsequent comparison of gene expression based on RNA-Seq at 24, 48, 72 and 96 hpi with that of mock (non-inoculated) samples showed that 3,091 disease responsive genes. Among them, 477 were differentially expressed genes (DEGs) (fold change >2, Padj < 0.05) between the resistant and susceptible RILs. Based on expression profiling, these DEGs were clustered into six expression clusters (C1-C6). In Cluster C1, 56 genes were up-regulated in the susceptible RIL whereas in C2, 79 genes were up-regulated in that RIL, mainly at 96 hpi. Cluster C3 contained 91 genes that were up-regulated in the resistant RIL LR-66-528 at 24, 72 and 96 hpi. A total of 97 genes in C4 were significantly up-regulated in LR-66-524 at 24 and 48 hpi. Cluster C5 with 51 genes was the smallest cluster with genes up-regulated in the resistant LR-66-528 and down-regulated in the susceptible LR-66-524, as were 95 genes in Cluster C6. DEGs were functionally annotated to identify those with known functions in disease resistance proteins, such as LRR and NB-ARC domain disease resistance protein, Protein Detoxification, LRR receptor-like kinase family proteins, and Wall-associated Ser/Thr Kinases. The expression of 21 of these genes was validated using RT-qPCR, which confirmed up- or down-regulation as in the RNA-Seq data. Comparison of DEGs and genes in QTLs associated with resistance to anthracnose revealed that nine DEGs were located in the resistance QTL region of chromosome 2, ten in the QTL region of chromosome 5 and three in the QTL region of chromosome 7 of L. ervoides. The identified candidate genes associated with resistance should be valuable targets for the future gene function analyses.

Description

Keywords

RNA-Seq, Lens ervoides, Anthracnose, Colletotrichum lentis, Disease resistance

Citation

Degree

Master of Science (M.Sc.)

Department

Plant Sciences

Program

Plant Sciences

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