The study of tolerance mechanisms of higher plants against abiotic stress conditions to improve the production of crop plants is a very important research area in the world. In this project we will use yeast to identify genes of Selaginella lepidophylla th at are important for its exceptional drought tolerance.
This plant is able to survive in a complete dry form for many years. One of the reasons for its high stress tolerance is the very high trehalose levels found in this plant. Trehalose is well-known for its stress protection characteristics. It is synthesised in two steps. Tps1 catalyzes the formation of trehalose-6-phosphate, which is then converted to trehalose by the activity of the Tps2 enzyme.
Previously we have isolated the S. lepidophylla TPS1 gene . Expression of this gene in a yeast tps1 mutant did not complemented the deletion phenotype (growth on glucose and trehalose synthesis). N-terminal truncation of this gene, however, resulted in an allele with very high activity upon expression in yeast, indicating N-terminal inhibition on the activity of the rest of the protein.
We have constructed a S. lepidophylla cDNA library in a yeast expression vector and we will use this library to screen for genes that may interact with the N-terminal sequence of the previously characterized SlTPS1 gene by selection of transformants that grow on glucose and that produce high levels of trehalose. So far, no real TPS2-like genes have been isolated from plants.
As S. lepidophylla accumulates high levels of trehalose, we expect a functional TPS2 enzyme to be present. We will use the library to transform the temperature sensitive yeast tps2 strain and screen for transformants that allow growth at 39 °C.
Finally we will also identify other components that may increase stress tolerance of yeast cells. The components resulting in the best stress tolerance in yeast cells will also be transformed into A. thaliana and. tested for stress tolerance.
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