14 ERA-CAPS: INvestigating TRiticeae EPIgenomes for Domestication (INTREPID)

Lead Research Organisation: University of Liverpool
Department Name: Institute of Integrative Biology


Plant breeding uses DNA sequence variation to make new allelic combinations for crop improvement. Our creation of the first wheat gene sequence assemblies (Brenchley et al Nature 491, 705) has enabled new levels of high throughput precise genotyping for breeding this globally important crop. Nevertheless, there are other levels of heritable variation, such as epigenetic modifications, that are widely thought to play a key role in shaping genomes and creating new variation. We have recently developed highly efficient re-sequencing technologies for wheat that can measure DNA methylation in genes of multiple lines. This provides an outstanding opportunity to assess epigenetic variation in a major polyploid crop and understand how it may influence traits. The overall objective of this proposal is to use newly available wheat genome resources, together with our innovative application of exome capture and bisulphite sequencing, to measure epigenetic modifications in wheat genes, and relate these to gene expression and the acquisition of new phenotypes, and how they may contribute to genetic changes such as gene loss during polyploid formation.

Technical Summary

The production of new hybrids is an important way of improving crops as they exhibit novel traits directly after hybrid formation, which are not found in progenitor parents. Growing evidence points to possible epigenetic origins for these emergent phenotypes. The scale and heritability of epigenetic modifications therefore needs to be measured, related to potential changes in gene and chromosome function and then taken into account in breeding as a source of variation in breeding.
Here, we aim to build on our collective experience in plant epigenetics and genomics to map the epigenome of bread wheat. Outputs of this project will be of immediate value for breeders for understanding the extent and contribution of epi-allelic variation to traits and in the choice of parental epi-allelic variation in making new hybrids. The project will also exploit experimental advantages of wheat to understand how epigenetic marks are re-programmed during the formation of new wheat hybrids, and how their independently maintained genomes influence each other during stabilization of the new hexaploid genomes. We have established four key foundations for mapping and understanding the wheat epigenome: the first genome sequence assembly of wheat; an efficient method for the cost-effective sequencing of the gene space of multiple wheat genomes and for determining genome-wide DNA methylation patterns an improved understanding of the mechanisms of epigenetic inheritance and evidence of altered gene expression in wheat hybrids.
This will generate new knowledge of how epi-alleles are formed and maintained, how the genomes of polyploid wheat influence each other, and how they influence gene function. It will have an important impact on wheat breeding by establishing the extent of epigenetic variation in wheat lines and its consequences on genome function and predicted phenotypes. Such information can guide the choice of parents for hybrid formation and explain aspects of missing heritability.

Planned Impact

Wheat is one of the three global crops providing the bulk of human nutrition. Large-scale coordinated research programmes are aiming to increase yields and reduce agricultural inputs in order to meet future consumption patterns and to mitigate the predicted effects of a changed growing climate. This project will have a fundamentally important impact on wheat breeding by understanding, for the first time, the extent of inherited epigenetic variation in wheat lines, and its consequences on genome function and predicted phenotypes. Such variation has been predicted to contribute to creating new phenotypes and heterotic yield increases in hybrids, but it has not yet been assessed at a genome scale in wheat. Such variation may explain aspects of missing or low heritability and could be used in breeding programmes. The project will show how new epigenetic variation is generated in hybrids, how such variation is stabilized, how new patterns of gene expression are created and stabilized; and how patterns of epi-alleles and traits can be influenced by the environment. This can guide crop improvement strategies by determining the choice of parents for hybrid formation, and by identifying subsequent epigenetic variation and measuring its stability across generations before incorporation into breeding programmes. In addition to these basic and applied outputs of this project, we will develop bioinformatic tools for identifying and analysing epigenetic marks and tracking their potential phenotypic consequences through gene expression network analyses. These impacts will be delivered through the open cyber-infrastructure provided by iPlant and through databases such as Ensembl genomes, through close engagement with breeders, and through publications and training programmes.


10 25 50

Related Projects

Project Reference Relationship Related To Start End Award Value
BB/N005104/1 01/10/2015 30/09/2016 £425,122
BB/N005104/2 Transfer BB/N005104/1 01/10/2016 30/09/2018 £309,763
Description First description of the wheat epigenome:
• Developed technology/methodology advance combining sequence capture, bisulfite treatment and genome specific assignment of reads using homoeologous SNPs
• Analysed genome wide methylation patterns including reports of genome specific methylation and conserved methylation patterns across the three genomes
• Associating methylation of a single genome with genome specific gene expression
• shown Non-CpG methylation is enriched in sub-genome specific methylation suggesting a potential association of sub-genome specific methylation with pseudo genes
• Shown that tri-genome methylation is highly conserved with a diploid wheat progenitor while sub-genome specific methylation shows more variation i.e. context specific variation.

Population epigenomic diversity across the A. E. Watkins bread wheat landrace cultivar collection

There is a high level of genetic diversity in the A. E. Watkins bread wheat landrace collection and, we predict, also a high level of epigenetic diversity. This represents a dual source of variation not used in modern breeding that can be exploited for wheat improvement. Here, we analyze genotype and DNA methylation, an important mechanism of epigenetic gene expression control that can be passed between generations, across a core set of the Watkins collection. This core set includes 105 lines and captures the majority of the diversity in the Watkins collection. Using sodium bisulfite treatment and targeted gene enrichment we can survey genome-wide methylation and genotype across the three sub-genomes of allohexaploid wheat.

Using a 12Mb capture probe set we were able to analyze an average of 48 Mb of the wheat genome per sample at a minimum depth of 10X. Methylation shows high variability within the Watkins collection with 42.1% of analyzed sites classified as single methylation polymorphism sites, or SMPs, between the samples. The vast majority of these SMPs (91.5%) are rare variants that were seen in less than 10% of the samples. Tri-genome methylation is more conserved between accessions than genome specific methylation and therefore likely to be the most stable form of methylation. Genome specific methylation sites show enrichment for homoeologous SNPs that differentiate the genome that is methylated from the other two sub-genomes and this SNP typically infers a CpG site from a non-CpG site.

We have observed that methylation is a standalone source of variation in the absence of genetic variation, however, it is clear that if two wheat accessions show more closely related genotypes then their methylomes are more likely to be related. Both methylation and genotype are clearly influenced by the geographical origin of the sample, although it appears that genotypic profiles cluster across wider geographic regions while the methylation profiles of accessions tend to cluster into more local groups. These lines clustering locally by methylation may be more likely to have evolved with similar environmental conditions for growth and this may have influenced their methylation profiles. Therefore, we hypothesize that methylation acts as a fast-adaptive response to environmental stimulus that may later become 'hard coded' as SNPs.

We are currently collaborating with Klaus Mayer's team in Munich to assess the gene networks that are targeted by methylation across the Watkins collection and how these vary between accessions. Furthermore, we are analyzing RNA-seq data for 12 of the samples from the Watkins collection to allow correlation of genotype, methylation and gene expression in this analysis. Finally, we are working with Cold Spring Harbor laboratories to sequence the full methylome of Chinese Spring wheat.
Exploitation Route This was the first set of objectives of our grant we are now moving forward.
have now complete the second set of objectives and are working on the last two.
Sectors Agriculture, Food and Drink
Description INvestigating TRiticeae EPIgenomes for Domestication 
Organisation Cold Spring Harbor Laboratory (CSHL)
Country United States of America 
Sector Charity/Non Profit 
PI Contribution Development of approaches to sequence the epigenome and bioinformatics pipelines
Collaborator Contribution access to material and expertise in systems genomic and bioinformatics.
Impact not yet
Start Year 2015
Description INvestigating TRiticeae EPIgenomes for Domestication 
Organisation Helmholtz Association of German Research Centres
Department Helmholtz Zentrum München
Country Germany, Federal Republic of 
Sector Public 
PI Contribution Development of approaches to sequence the epigenome and bioinformatics pipelines
Collaborator Contribution access to material and expertise in systems genomic and bioinformatics.
Impact not yet
Start Year 2015