Last November, the 10+ Genome Project and its international team of researchers were hailed for their role in sequencing the genomes of 15 wheat varieties from around the world. It was an astounding accomplishment with great potential for cereal breeding, particularly for a sector that struggles to keep up with the investments being made in corn or soybeans.
Now, that research is being fed into the 4DWheat project, which aims to apply the latest genomic strategies to boost yields and reduce disease losses.
The 4DWheat project gets its name because its goal is “harnessing Diversity, advancing Domestication, enabling Discovery and expediting Delivery” in order to produce new sources of genetic variation. The project’s timeline is 2019 to 2024, with $11.2 million in funding.
“Domestication of exotic wheat is one of the five major activities of this project,” says Dr. Sateesh Kagale, associate research officer with the National Research Council Canada (NRCC) in Saskatoon. “In collaboration with Dr. Curtis Pozniak at University of Saskatchewan, we conceptualized a genome-editing strategy technology to reveal the hidden breeding value of wild relatives of wheat.”
Kagale stresses that the use of gene editing is a complement to conventional breeding efforts, not a replacement. CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) technology is also opening many doors to the discovery and use of genetic diversity, for instance by “fixing” mutations that create adaptive traits. Compared to selective breeding, it offers a chance to create major changes at significantly lower cost.
Among its many applications, gene editing enables a faster and more efficient conversion of alleles once considered undesirable into desirable ones, and using CRISPR/Cas-based technologies can result in a range of DNA edits synonymous to those found in natural populations.
That’s a huge asset for cereal breeding which hasn’t seen the same levels of investment as corn, soybeans or canola, mostly because of the complexity of the wheat genome and regulatory barriers. It also means more potential for incorporating desirable traits from so-called wild relatives.
“Genetic variation is the most essential component for any breeding program to develop crop varieties with improved yield, quality and disease resistance,” says Kagale. “Wild relatives are a rich reservoir of genetic variation, but it’s difficult to assess their actual breeding value. Our project will develop methods to assess breeding value of wild relatives which will facilitate incorporation of individuals with the highest value into a breeding program.”
Gene editing is a more precise editing process, allowing researchers and breeders to target specific genomic regions to enable them introduce new genetic variation faster and more efficiently.
“Considering the controversies and public concerns regarding GM wheat and cereals, use of gene editing to create additional genetic variation in elite or exotic wheat becomes critical,” says Kagale.
“Although Canada has taken a step towards exemption of gene-edited organisms from safety assessments by Health Canada and the Canadian Food Inspection Agency (CFIA), global attitudes towards the fine nuances of gene editing will have huge impacts on how this technology is implemented and traded.”
One Agriculture and Agri-Food Canada (AAFC) researcher likes to describe CRISPR by saying it helps breeders “correct a spelling mistake” in the plant DNA, and Kagale agrees this is an effective analogy. However, the spelling mistake metaphor is only one application in a widely expanding array of potential genomic capabilities. Researchers can also use CRISPR to conduct DNA imaging in live cells, or to initiate epigenetic changes, among other applications.
“The success of CRISPR technology is related to its high precision, ease of design and lower cost compared to other gene-editing tools,” says Kagale. He cites RNAi (ribonucleic acid interference) as an older method which is still in use among researchers. “CRISPR/Cas-based technology is rapidly expanding and improvements are being seen in specificity, precision and off-target effects, editing capabilities and ease of use in target organisms.”
Of particular note is the development of Cas9 novel variants, Cas9 orthologs — genes in different species that evolved from a common ancestral gene — and CRISPR-associated enzymes. The development of novel Cas9 variants has enabled the manipulation of gene expression and targeted modification of genes, all without “breaking” DNA.
“Where possible, we try to mimic natural alleles of the target genes to achieve a desired phenotype, which will help us in avoiding regulatory hurdles,” adds Kagale, noting wheat dwarfing alleles Rht1 and Rht2 as examples. “Each of these contains a single base-pair mutation that results in reduced plant height. There are many genes associated with yield, grain quality, herbicide tolerance and nitrogen-use efficiency in which single base-pair mutations can convert undesirable phenotypes into desirable ones.”
Raising the bar
Kagale notes the recent sequencing of multiple wheat genomes has laid a solid foundation for the application of breeding technologies such as genomic selection and genome editing for crop improvements. More than that, these new resources have put wheat on par with corn, soybeans and canola in terms of crop innovation, although it will take a few years to see actual benefits. It will also require significant investment in wheat research because of the complexity and size of the wheat genome.
The ability to incorporate desirable genetic traits from wild relatives is also an important factor, as seen in Pozniak’s work in the 10+ Genome Project. Mimicry of natural alleles in target genes helps avoid regulatory hurdles but also incorporates needed genetic variation. In elite breeding lines, that characteristic has been molded by domestication and plant selection.
For instance, in wheat, plant height, photoperiod, vernalization response and grain threshability are domestication-related traits. Yet the undesirable alleles of those traits in wild relatives of wheat masks their agronomic potential.
To put it all in perspective, more than 500,000 accessions (groups of related plant material) of wheat relatives are contained in gene banks around the world, yet less than 10 per cent of genetic variation in landraces and wild relatives of wheat has been incorporated in modern varieties.
“In the 4DWheat project, we’re using CRISPR technology to fix the allele modifications required to recreate domestication traits in exotic germplasm,” says Kagale. “Wild relatives of wheat are not adapted to Canadian conditions, and genetic barriers such as sequence divergence and strict regulation of chromosome pairing affect their utilization in conventional, selective breeding.”
A "can't miss" approach
In the course of an eight- to 12-year breeding process, new varieties can be developed that exhibit desirable traits such as disease tolerance or high yields. But unanticipated changes in one trait can have an impact on classification options or market viability of a variety.
Unlike corn or soybeans, registration of wheat varieties is overseen by downstream millers and processors.
If a new variety fails to meet their specifications, it will not be registered. Now, those lines that are missing a key characteristic — the “near misses” — have the potential to reach growers, thanks to these CRISPR innovations.
The 4DWheat project is led by Dr. Curtis Pozniak (University of Saskatchewan) and Dr. Sylvie Cloutier (Agriculture and Agri-Food Canada). It has been made possible through funding from Genome Canada, Agriculture and Agri-Food Canada (AAFC) Partnership, Canadian Wheat Research Coalition, Western Grains Research Foundation, Illumina, Saskatchewan Wheat Development Commission, Alberta Wheat Development Commission, Manitoba Wheat and Barley Development Commission, Viterra, Province of Saskatchewan, and Ontario Ministry of Economic Development, Job Creation and Trade. Genome Prairie is providing administrative support.