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Exploring barley mutants for molecular dissection of chlorophyll biosynthetic enzymes and chloroplast development
Project
Project code: DFG-411988294
Contract period: 01.01.2018
- 31.12.2020
Purpose of research: Experimental development
Mutation research became a hot topic exactly 90 years ago. Barley chlorophyll mutants were used as a tool in the mutant studies since their phenotype is obvious already at the seedling state; mutants not able to make chlorophyll were easily spotted as yellow or white plants, which indicated a successful mutagen treatment. These chlorophyll mutants have been kept ever since and provide today a fantastic opportunity to reveal the molecular mechanism of chlorophyll biosynthesis and chloroplast development. The mutations are lethal since the plants cannot survive without chlorophyll but due to the energy rich seeds of barley the chlorophyll-less mutants can anyway grow to a seedling of 10 cm, which allow analyses of the plants that cannot be performed with Arabidopsis mutants. The long-term goal of this study is to reveal the genes and deepen the understanding of the corresponding enzymes and proteins participating in chlorophyll biosynthesis and affecting chloroplast development. Thus, in the Hansson lab, different subprojects are at different stages along a gene-identification-to-protein-characterization scale and I will participate in projects at different stages. I will perform gene identification to identify the totally unknown genes Xantha-a and Xantha-m at the DNA level using xantha-a and xantha-m mutants which will be analyzed by genotyping-by-sequencing. Candidate genes located in the identified region will be sequenced from the 15 and 5, respectively, available allelic mutants. A gene with mutations in the mutant lines suggests that the correct gene has been identified. The gene will be cloned and used in expression systems of Escherichia coli in order to produce Xantha-a and Xantha-m proteins for biochemical assays and further characterization. Barley genes of Mg-chelatase and cyclase have previously been identified by members of the Hansson lab and expression systems have been developed. Thus, the studies of these enzymes are more directed to understand their mechanisms. Since the design of the proteins reveal much of their functions, the Hansson lab have focused on 3D structural studies employing X-ray crystallography and cryo-electron microscopy. I am very excited to take part in these experiments since that will broaden my repertoire of molecular techniques.
Section overview
Subjects
- Biotechnology
