Scientific achievements
Chromosome associations and recombination between H. chilense and wheat have been promoted for the first time in the background of ph1 mutant in the context of this project.
A 4Hch substitution line in durum wheat has been developed as a required breeding tool to transfer resistance to Septoria tritice into durum wheat.
Inter-specific recombination between chromosomes 4Hch and 7Hch and their homoeologous wheat chromosomes has been promoted in the background of the ph1 mutant.
A set of wheat lines carrying small introgressions for chromosomes 4Hch and 7Hch has been obtained.
Wheat lines with increased carotene content have been developed. These new wheat-H. chilense translocation lines can be used as a powerful tool in wheat breeding programs to enrich the diet in bioactive compounds.
Genes conferring resistance to Septoria tritice have been narrowed to a 4Hch chromosome segment.
Proteins involved in chromosome pairing during meiosis have been identified using rice as model species.
Meiotic replication has been studied in wheat. The effect of the Ph1 locus on meiotic replication has been assessed.
Novel and/or unconventional methodologies
The development of a methodology to isolate enough amounts of meiocytes from anthers in each stage of the meiosis for cytogenetics, proteomics or flow cytometry analysis was a challenge and could be consider as a technological achievement (see Collado-Romero et al, 2014). Meiocytes are a small number of cells within the anther. Having meiocytes isolated from the rest of the anther tissues makes possible to process specifically samples for meiosis by flow cytometry, mass spectrometry or real-time PCR, to determine, for example, when replication occurs, identify specific meiotic proteins or quantify the level of expression of a specific gene in a particular stage of meiosis. This approach allow the analysis of the samples with a minimal level of DNA/proteins contamination from other anther tissues, optimizing the results. In fact, proteins involved in meiosis are a small proportion of total anther proteins and will be practically undetectable in case of using whole anthers for mass-spectrometry analysis. Similarly, the expression of a particular meiotic gene can be measure more accurately using the approach developed in this project. This methodology can be applied to other plant species.
In addition, an efficient and user-friendly method for the screening of small random genomic introgressions from Hordeum chilense in the wheat germplasm has been developed in this project (see Rey et al, 2017). DNA from wheat lines carrying putatively H. chilense introgressions was immobilized on a nylon membrane and blocked with bread wheat genomic DNA. The detection of the alien genomic introgressions was carried out using total genomic DNA from the alien species as a probe, labeled with biotin-11-dUTP. This technique allowed a rapid and reliable screening for detecting small random H. chilense genomic introgressions introduced during a breeding program in the background of bread wheat when molecular markers cannot be used and the in situ hybridization makes the detection process very expensive and high time consuming. Thus, using this dot blot screening, the number of plants required for a deeper analysis by molecular markers or in situ hybridization decreased, saving time and money. The methodology is also suitable to detect other wheat related species such as Hordeum vulgare, Secale cereale and Agropyron cristatum in the wheat background.
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