Dissection of the genetic control of drought tolerance in red clover using transitional genomics

Supervisors: Dr Leif Skot, Dr Michael Abberton, Dr Glyn Jenkins

Abiotic stresses such as drought, salt, and extremes of temperature can severely limit the productivity of crop plants. Climate change is likely to lead to increases in both the severity and frequency of occurrence of these conditions. Breeding for improved tolerance to these stresses is thus increasingly important in order to maintain crop productivity.
The proposed work will focus on the genetic dissection of drought stress in red clover (Trifolium pratense) an increasingly important forage crop in the sustainable agriculture of temperate agriculture. It will complement  work  being conducted under recently awarded  ERANET-Plant Genomics funding, one objective of which is to anchor the physical map of red clover to the genome of the model legume Medicago truncatula (barrel medic), and align a clover genetic map to the physical map. The ERANET study will include extensive BAC end sequencing (Arizona Genome Institute) cytogenetic analysis using FISH and GISH (Wageningen) and a considerable bioinformatic component (MIPS in Munich). IGER will co-ordinate this work and be responsible for genetic mapping and alignment of physical and genetic maps.
Recent work in the Legume Breeding and Genetics (LBG) Team at IGER has utilised the recently available M. truncatula Affymetrix gene chips to identify genes which are up regulated in both M. truncatula and clover in response to drought stress. Candidate genes are currently being confirmed by RT-PCR partly by a PhD student within the team. A red clover genetic map has been produced in the work of another PhD student within LBG. Take together these resources represent a considerable platform for dissection of the genetic basis of key traits including cloning of QTL

Objectives

Identify cross orthologous genes in red clover, which have been shown to be involved in the response to drought stress. This will be achieved by utilising the A. thaliana and M. truncatula genomics resources and databases (such as www.arabidopsis.org, www.medicago.org and www.tigr.org), in combination with the sequencing information that becomes available in red clover from the ERANET-PG project. This work will also include analysis of those genes identified by the Affymetrix gene chip experiments and will provide the student with bioinformatics skills.
Utilising the EST database from red clover to identify cross orthologous sequences (COS) by searching against databases of A. thaliana, Lotus japonicus and M. truncatula. This will increase the number of genes available for subsequent mapping in red clover, and further improve the bioinformatics skills of the student.
Identify single nucleotide polymorphisms (SNP) using DNA sequencing and/or single strand conformational polymorphism, between parents of a mapping family in the drought-related genes as well as the general COS sequences. A red clover F1 mapping family is available for mapping of genetic markers.  Mapping of the former group of genes will enable identification of chromosomal regions containing functionally related genes, and explore possible syntenic relationships with the model legumes. Such syntenic relationships can be further explored by mapping of the EST-based COS markers. The information that will be acquired in this project will also directly benefit the ERANET-PG project by allowing the gene-specific SNP markers to be genetically mapped, and thus aligned with the physical map.
Co-location of candidate gene position with drought tolerance QTL. There is extensive experience within LBG with regard to the phenotyping of the morphological and physiological characteristic of the response to drought stress.
Cloning of drought tolerance QTL
Transcript analysis (functional testing) of candidate genes by QPCR in drought and undroughted material

The project will equip the student with a range of relevant skills for the post-genomics era. These include: marker development (SNPs), RT-PCR, comparative mapping, in silico approaches to marker design and comparison between genomes, drought phenotyping and physiological analysis, QTL mapping. The student will be involved in the process of model to crop translational genomics that is key to the use of modern approaches in crop involvement. The work will combine experimentation in laboratory, glasshouse and field with bioinformatics. He/she will gain considerably from other work going on within the team and particularly in the international collaboration under ERANET-PG.