Abstract
Rye is used as food, feed, and for bioenergy production and remain an essential grain crop for cool temperate zones in marginal soils. Ergot is known to cause severe problems in cross-pollinated rye by contamination of harvested grains. The molecular response of the underlying mechanisms of this disease is still poorly understood due to the complex infection pattern. RNA sequencing can provide astonishing details about the transcriptional landscape, hence we employed a transcriptomic approach to identify genes in the underlying mechanism of ergot infection in rye. In this study, we generated de novo assemblies from twelve biological samples of two rye hybrids with identified contrasting phenotypic responses to ergot infection. The final transcriptome of ergot susceptible (DH372) and moderately ergot resistant (Helltop) hybrids contain 208,690 and 192,116 contigs, respectively. By applying the BUSCO pipeline, we confirmed that these transcriptome assemblies contain more than 90% of gene representation of the available orthologue groups at Virdiplantae odb10. We employed a de novo assembled and the draft reference genome of rye to count the differentially expressed genes (DEGs) between the two hybrids with and without inoculation. The gene expression comparisons revealed that 228 genes were linked to ergot infection in both hybrids. The genome ontology enrichment analysis of DEGs associated them with metabolic processes, hydrolase activity, pectinesterase activity, cell wall modification, pollen development and pollen wall assembly. In addition, gene set enrichment analysis of DEGs linked them to cell wall modification and pectinesterase activity. These results suggest that a combination of different pathways, particularly cell wall modification and pectinesterase activity contribute to the underlying mechanism that might lead to resistance against ergot in rye. Our results may pave the way to select genetic material to improve resistance against ergot through better understanding of the mechanism of ergot infection at molecular level. Furthermore, the sequence data and de novo assemblies are valuable as scientific resources for future studies in rye.
Highlights
Rye (Secale cereale L.) is a widely popular cereal crop, especially in northern Europe, due to its high nutritional value and tolerance to unfavorable environmental conditions
We presented a stringent workflow in this study for processing of RNAseq data, obtaining de novo assembly, assessment of assembly, functional annotation and differential gene expression from spikes of the non-model plant S. cereale
We identified the significantly enriched gene ontology (GO) terms of these differentially expressed genes (DEGs) that were significantly associated to carbohydrate metabolic processes, hydrolase activity, pectinesterase activity, cell wall modification, pollen development and pollen wall assembly (Fig. 6)
Summary
Rye (Secale cereale L.) is a widely popular cereal crop, especially in northern Europe, due to its high nutritional value and tolerance to unfavorable environmental conditions. We presented a stringent workflow in this study for processing of RNAseq data, obtaining de novo assembly, assessment of assembly, functional annotation and differential gene expression from spikes of the non-model plant S. cereale In this Genomic era, knowledge about the ergot disease has enhanced considerably but the fundamental underlying mechanisms are still poorly understood. RNA sequencing is a highly efficient method, but false positives still occur due to sensitivity of this technology Keeping this drawback in mind, we used both a de novo assembled and the draft reference genome of rye to count the differentially expressed genes in a moderately ergot resistant compared to an ergot susceptible hybrid with and without inoculation. To the best of our knowledge, this is the first attempt with proper experimental considerations to unravel genes involving in ergot infection mechanism in rye hybrid
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