Epigenetic changes in barley in response to salinity stress

Author: Jason Smith

Smith, Jason, 2020 Epigenetic changes in barley in response to salinity stress, Flinders University, College of Science and Engineering

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Epigenetic mechanisms are potentially heritable molecular changes that affect gene expression, leading to differences in phenotype without changing the DNA sequence of the organism. In plants, such mechanisms are involved in the control of a range of processes, including response to stress. DNA methylation is an epigenetic mechanism used by organisms to adapt to changing environmental conditions by altering localised accessibility of the genome to transcription factors, thus ultimately affecting gene expression levels. The sequence and organ specificity of this stress induced de-novo DNA methylation in plants is guided by a class of small RNAs (sRNAs), typically 23 to 24 nt long.

This project uses epiGBS, a reduced representation genome bisulphite sequencing method, coupled with small RNA and whole transcriptome Next Generation Sequencing to investigate interactions between sRNAs, DNA methylation and gene expression in the leaves and roots of barley under salt stress. EpiGBS was chosen as a method for gaining information about the methylation states of the genome since the barley genome is large, at 5.1 Gbp, making whole genome sequencing prohibitive. The method reduces the complexity of the genome by restriction enzyme fragmentation and sequencing based on fragment size selection, with genome coverage scalable by size selection and number of reads. Hordeum vulgare (barley) was chosen for this study as it is the fifth most important crop in global agriculture with good coverage of genome sequence information. It is anticipated that findings in this research should be applicable to other cereal crops such as wheat.

Analysis of small RNA sequence data identified 59 new H. vulgare microRNAs (miRNAs) and corresponding precursor hairpin-loop sequences found that have not been previously reported in miRBase. Of these newly discovered miRNAs, 44 did not have sequence similarity to any previously identified miRNAs in other plant species and 15 were similar to known miRNAs in other plant species. Eight of these newly discovered miRNAs correlated with salinity stress and are likely to be involved in stress response. DNA methylation changes were found in response to salinity stress, with 1,210 loci in leaf tissue and 513 loci in root tissue found corresponding to a change in 23/24 nt sRNA expression that targets a protein coding gene. The linkages between sequence information from DNA methylation, small RNA and the transcriptome will lead to a greater understanding of how this crop deals with this important stress, and provide an extremely useful avenue for further research.

Keywords: epigenetic modification, DNA methylation, sRNA, barley

Subject: Biotechnology thesis

Thesis type: Doctor of Philosophy
Completed: 2020
School: College of Science and Engineering
Supervisor: Peter Anderson