Periodic Reporting for period 1 - AdaptiveTarget (AdaptiveTarget: Accessing haplotype variation at complex loci with optimized targeting and adaptive sampling)
Reporting period: 2024-01-01 to 2025-06-30
Genetic variation at resistance loci is crucial for sustained crop yield, given the challenges and increased pest and pathogen pressures resulting from a warming climate. Despite recent advances in massively parallel sequencing, disease resistance loci and other complex genomic regions of great practical, economical and scientific importance remain challenging to study. The severe limitations of current methods pose a major problem for fully understanding and harnessing genetic variation for crop improvement. We evaluated AdaptiveTarget as a bioinformatic method based on targeted long-read sequencing, to allow researchers and breeders to obtain population-level haplotype information for complex genomic regions such as resistance genes and self-incompatibility loci in plants. To explore the innovation potential of our ERC-funded research we first aimed to undertake research to test and validate the project idea. Validation benefits from known haplotype data for the complex S-locus supergene, produced in the ERC-funded project SuperGenE. To demonstrate the general utility of the method we tested it on a real-world example of complex loci of agronomic importance. We also developed a piece of software that we aim to release after completion of the project.Together these experiments provide a comprehensive assessment of bioinformatically targeted long-read sequencing for resolving complex genomic regions in plants. By evaluating both the technical performance and biological utility of this method, our project highlights its potential as a powerful tool for plant genomics, with implications for evolutionary biology, gene discovery, and crop improvement.
In this project, we have tested and optimized targeted long-read sequencing of complex loci in three plant systems: (1) the self-incompatibility S-locus in Linum species, (2) a complex disease resistance locus in a wild wheat relative, and (3) the S-locus in Brassicaceae species. Each of these systems presents distinct challenges and use cases, from targeting across species boundaries to multiplexing multiple samples within a single sequencing run.
We first tested the validity and limitations of our chosen approach for targeted sequencing using sequences of the Linum distyly S-locus generated in our ERC project SuperGenE as ground truth. At this stage, we identified key limitations in the planned approach, which prompted us to further develop and optimize our method. Second, we tested the modified method on Linum species with varying lengths of their S-loci. Third, we developed a new piece of software and demonstrated the functionality of targeted long-read sequencing together with our software in two real-world examples: the sequencing of a complex resistance gene in a bread wheat relative, and the sequencing of the S-locus in the Brassicaceae. By evaluating both the technical performance and biological utility of this method, and by developing custom software, the project will facilitate the use of this powerful tool in plant genomics.
We first tested the validity and limitations of our chosen approach for targeted sequencing using sequences of the Linum distyly S-locus generated in our ERC project SuperGenE as ground truth. At this stage, we identified key limitations in the planned approach, which prompted us to further develop and optimize our method. Second, we tested the modified method on Linum species with varying lengths of their S-loci. Third, we developed a new piece of software and demonstrated the functionality of targeted long-read sequencing together with our software in two real-world examples: the sequencing of a complex resistance gene in a bread wheat relative, and the sequencing of the S-locus in the Brassicaceae. By evaluating both the technical performance and biological utility of this method, and by developing custom software, the project will facilitate the use of this powerful tool in plant genomics.
In this project, we tested the validity and limitations of a new bioinformatic approach to targeted long-read sequencing of complex loci of major biological and agricultural importance. We identified and developed strategies for mitigating limitations of the method, and developed new software to apply it. Finally, we demonstrated its utility for studying complex loci of interest for both fundamental and applied purposes. We have presented the results to a diverse set of target audiences and investigated avenues to licensing and commercialization. To ensure further uptake and success, we plan to release the software we developed. This project will therefore help enable the use of targeted long-read sequencing in both fundamental and applied settings.