Molecular mechanisms of drought stress tolerance and water use efficiency in the monocot crops Sorghum and rice

The general academic aims of this project are to understand the fundamental underlying processes in drought stress tolerance and water use efficiency, using the staygreen Sorghum lines as a model. The general strategic aims of this project are to transfer this knowledge into another monocot crop, rice. To achieve this the specific objectives of this project are:
(1) Testing which specific biochemical, cellular, developmental and molecular pathways (based on gene expression patterns) are responsible for the staygreen agricultural trait
(2) Identification of genes which control these processes e.g. we have identified a transcription factor and an E3 ubiquitin ligase which are hypothesised to control drought responses
(3) Identification of orthologous genes in rice, the worlds most important crop
(4) Creating transgenic rice lines in which these pathways are modified to emulate the sorghum staygreen condition, and testing these lines for improved drought tolerance/WUE.
Methodology
We have produced transgenic sorghum lines to test two genes which we already hypothesise are responsible for the staygreen phenotype in sorghum. By changing the expression of these in non-staygreen sorghum backgrounds, we will test for the acquisition of the staygreen trait. These lines will thus be tested for drought tolerance and water use efficiency, using for instance, measurements of Fv/Fm and stomatal conductivity
Other genes will be selected from analysis of our existing transcriptomic data, orthologues will be cloned from rice, and constructs made to either overexpress, or suppress expression in rice and transgenic lines produced. These will be tested for drought tolerance and water use efficiency as described above. Where appropriate, testing function of cloned genes in transgenic Arabidopsis may be appropriate as a quick way to determine/confirm function, and therefore allow the screening of even more genes.
Timetable of Activities
Year 1: Testing transgenic sorghum lines for altered drought tolerance, WUE, developmental aspects, senescence, production of compatible solutes and expression of stress genes. Analysis of staygreen transcriptomic data, identification of candidate genes for analysis in rice;
Year 2: Cloning of rice orthologues, production of overexpression/suppression binary constructs, transformation into rice. Possible transformations into Arabidopsis (with other bespoke constructs). Characterisation of Arabidopsis lines
Year 3: Characterisation of transgenic rice lines, testing for lines for altered drought tolerance, WUE, developmental aspects, senescence, production of compatible solutes and expression of stress genes. Write thesis.
Novelty; No one has reported taking a transcriptomic approach to studying the staygreen traits of sorghum, and the candidates we have identified are novel, therefore academically this project will produce novel insights into plant drought tolerance and WUE, and strategically will provide novel pathways for breeding tolerance in crops.
Training to be received by student: Physiological measurements of drought stress, WUE, photosynthesis, gene expression measurements, realtime PCR, imaging reporter genes e.g. using luciferase, microarray analysis, recombinant DNA cloning, genetic transformation of bacteria, Arabidopsis and rice.