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Food Security Rotation Projects 2014-2015

Disease resistance and vaccine responses in chickens

Supervisor: Professor Jim Kaufman, Pathology

Project abstract: My primary interest is to understand the evolution of the adaptive immune system, but we have found that the most informative animal model has been the domestic chicken. So, we work on many aspects of the immune response in chickens, funded by the Wellcome Trust, BBSRC, animal vaccine companies and poultry breeding companies. At the moment, we have on-going projects on peptide binding of chicken class I molecules (Wallny et al 2006 PNAS, Koch et al 2007 Immunity), peptide transport by TAPs (Walker et al 2011 PNAS), NK cell receptors  (Rogers et al 2008 Semin Immunol, Rogers and Kaufman 2008 Immunogenetics), antigen presentation by class II molecules (Parker et al, in preparation), large scale typing of MHC molecules and relation to disease resistance in commercial and backyard chicken populations worldwide (Potts et al, in preparation), a highly polymorphic innate immune detection system called BG (Salomonsen et al in review at PLoS Genet), and genetics of response to in ovo vaccination (Butter et al 2013 Immunogenetics, Chan et al, in preparation), among other projects. Just starting up this fall is a BBSRC-supported project to develop a high through-put procedure for determining T cell epitopes in poultry pathogens.

Learning outcomes and skills acquired: For these projects, we use a wide variety of techniques taken from biochemistry, molecular biology, cell biology, immunochemistry and imaging, cellular immunology, genetics, genomics and population biology.  Depending on the project, the student would gain experience in polymerase chain reaction, cloning and sequencing, eukaryotic cell expression, protein purification, immunoprecipitation, western blot, vaccination of chickens, enzyme-linked immunosorbant analysis, cellular assays, flow cytometry, confocal microscopy, and whatever methods were appropriate to solve the problems at the time.

Project availability: I confirm my own availability for the duration of the project  (We cannot accept a project proposal for a term in which a PI is not available to supervise the student)

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Determining how Campylobacter jejuni grows and survives from ‘farm-to-fork’ and beyond...

Supervisor: Dr. Andrew Grant, Veterinary Medicine

Project abstract: Campylobacter jejuni is the leading cause of bacterial diarrhoeal disease worldwide, with faecal contamination of meat during processing a major route of transmission to humans. There is no vaccine, and measures aimed at reducing environmental contamination have so far proved ineffective.  We are determining the genes required by C. jejuni from farm-to-fork, with the aim of developing novel control and intervention strategies.    To enable a comprehensive study of mechanisms involved in growth/survival from farm-to-fork, we have constructed large random transposon mutant libraries in C. jejuni. We have/will screen these mutants through various in vivo and in vitro assays (e.g. colonisation of chickens and pigs; survival in flies, water and at low temperature; biofilm formation; adherence and invasion of cultured cells).  The relative abundance of mutants (input vs output) can be determined by massive parallel sequencing of transposon insertion sites (Tn-seq), to enable identification of genes that are specifically required under certain conditions.  Genes of interest will be selected and studied in detail.    Most of the Campylobacteraceae are helical, it is suggested that this shape is important for its ability to colonise its hosts and cause disease. To address this hypothesis it is essential to know how helical shape is determined, but we have limited understanding of this.  We have identified 3 peptidoglycan modifying enzymes which when mutated result in C. jejuni becoming rod-shaped, we are investigating the role of these proteins in helical cell shape formation, and how they may be manipulated for novel forms of intervention.

Learning outcomes and skills acquired: The project will involve functional genomics, bacteriology, molecular biology, microbiology, microscopy, sequencing and bioinformatics.  The student will be part of a dynamic group (including 5+ post-docs) working on related projects and will receive excellent support in a lab that is well funded and contains the equipment needed to conduct the research.  Lab meetings will be held weekly where the student will present and discuss data.  In addition, the student will attend a weekly meeting with ~30 other researchers, and weekly Departmental seminars, where they will get experience in presenting their work to a larger and multidisciplinary audience.

Project availability: Michaelmas and Lent Term

Other relevant themes: Basic bioscience underpinning health

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Does variation in nectar availability affect pollinator attention to crops?

Supervisor: Professor Beverley Glover, Plant Sciences

Second supervisor: Jane Thomas, (NIAB) National Institute of Agricultural Botany

Project abstract: An estimated 35% of global crop production depends on animal pollination, including key oil crops, legumes, fruit, fruit vegetables and nuts. However, a decline in pollinator numbers across the world has started to limit the odds of pollination and to reduce crop production rates [1,2]. Understanding which features of flowers attract pollinators and how those structures vary in crop plants is therefore a major goal in plant biology. Our group works on understanding the effects of changes in flower traits on pollinator behaviour, using bumblebees (Bombus terrestris) in controlled lab conditions [eg 3]. We also explore the molecular genetic control of the development of these structures [eg 4]. These approaches combine to give us insight into how key crop species can be improved to maximise food security in the face of pollinator decline. In this project you will explore the variability in nectar volume and sucrose concentration present across a range of Vicia faba (field bean) lines, to identify target lines for crop improvement. At the same time you will use bee behavioural approaches to assess the optimum combination of nectar volume and sucrose concentration for pollinator attraction and fruit set. Your data will feed directly into the breeding of Vicia faba to optimise yield under current limited pollination regimes, and provide a basis for future molecular genetic studies of nectar production.

References

[1] Basu, Bhattacharya, Ianetta. A decline in pollinator dependent vegetable crop productivity in India indicates pollination limitation and consequent agro-economic crises. Nature Precedings,hdl:10101/npre.2011.6044.1, 2011.

[2] Klein, Vaissiere, Cane, Steffan-Dewenter, Cunningham, Kremen, Tscharntke. Importance of pollinators in changing landscapes for world crops. Proc. Roy. Soc. B. 274:303-313, 2007.

[3] Whitney, Chittka, Bruce, Glover. Conical Epidermal Cells Allow Bees to Grip Flowers and Increase Foraging Efficiency. Current Biology 19, 1-6, 2009.

[4] Box, Dodsworth, Rudall, Bateman, Glover. Characterisation of Linaria KNOX genes suggests a role in petal spur development. Plant Journal 608, 703-714, 2011.

If you have other aspects of floral biology and pollinator attraction that you would like to investigate please contact Beverley to discuss project ideas.

Learning outcomes and skills acquired: This project will expose you to a combination of plant morphological and behavioural ecology skills. You will use standard techniques for nectar extraction and analysis of sugars. You may also use DNA/RNA manipulation techniques combined with microscopy to understand the production of nectar. You will use behavioural assays with caged bumblebees to understand how nectar concentration and volume influences pollinator foraging. You will develop skills in statistical analysis of morphological and behavioural data. It is very likely that your data will contribute to a research paper, giving you additional experience in scientific writing and the publication process.

Project availability: Michaelmas and Lent Term

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Discovering new genes involved in taste perception, with potential relevance to pest control

Supervisor: Professor. Chris Jiggins, Zoology

Second supervisor: Dr. Hannah Rowland, Zoology

Project abstract: This project unites researchers in genomics and behaviour at the University of Cambridge, and wildlife damage researchers at the United States Department of Agriculture, with the aim to identify the genes responsible for taste perception in European starlings – a pest species that is a major driver of current global habitat change, and a threat to the maintenance of global biodiversity, human health, and the success of human economic enterprises. Using RNASeq data, you will examine the starling chemosensory receptor transcriptome. You will identify and validate new taste gene transcripts. This will provide new discoveries in molecular biology and genetics, and will be an important advance toward understanding the molecular basis of avian sensory systems, and the design of non-harmful repellents. This is essential for the management of preferred habitats, and the reduction of economic impacts imposed by crop damage by Starlings. Starlings flocks can number 100,000 or more, and impact numerous agricultural crops including fruits, corn and sunflower. Flocks of starlings consume 2500 tonnes of livestock feed during a winter, and spoil more feed with their droppings. Yearly damage to agriculture from starlings totals $800 million in the USA alone. The most commonly used technique of managing starlings at feedlots and fruit production facilities is the use of taste aversive compounds that are bitter tasting. However, these compounds have not proven universally effective. Experts in vertebrate pest management have called for a greater understanding of the mechanisms underlying taste perception [1]. This project will address this request directly.    

Werner, S.J., and Clark, L. (2003). Understanding blackbird sensory systems and how repellent applications work USDA National Wildlife Research Center - Staff Publications Paper 287.

Learning outcomes and skills acquired: The student will gain experience in genomic sequence data analysis and manipulation including: learning a variety of clustering association and network techniques, and RNAseq sequence analysis techniques data. More generally they will be given a thorough grounding in project design and high throughput workflows.

Project availability: Michaelmas and Lent Term

Other relevant themes: World class underpinning bioscience

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Homologies in the evolution of shoot branching.

Supervisor: Dr. Jill Harrison, Plant Sciences

Second supervisor: Ottoline Leyser, Sainsbury laboratory

Project abstract: Branching is a key component of plant architecture and productivity as it determines how plants fill space in response to their environment. In flowering plants, a dominant shoot tip regulates the activity of shoot tips elsewhere in the plant to determine the final branching form via three plant hormones: auxin, cytokinin and strigolactone. The activity of these three cues is integrated via polar auxin transport pathways involving PIN-FORMED1 auxin transporters(1).  Unlike flowering plants, the earliest land plants had a single shoot, and the most ancient branching forms in the fossil record have a forked form(2). Such bifurcating forms are represented by modern lycophytes, which branch by amplification of the stem cells at the shoot tip followed by branch divergence during growth(3). Recent work in our lab has identified polar auxin transport via PIN transporters as a likely homology of branching pathways, but the point at which cytokinin and strigolactone were recruited in evolution to regulate branching is not clear.   This project will test the hypothesis that PIN-mediated auxin transport is a conserved regulator of branching using a combination of immunohistological, surgical and pharmacological approaches with plant hormones in the lycophyte Selaginella.   The results will be relevant to architectural engineering in crops as they will identify fundamental aspects of branching. 

1.   Domagalska & Leyser (2011). Nat Rev Mol Cell Biol 12:211-221. 

2.  Edwards et al. (2014). New Phytologist 202:50-78. 

3.  Harrison et al. (2007). Development 134:881-889.

Learning outcomes and skills acquired: The hormone treatments will involve plant tissue culture and micromanipulation, and Immunohistochemistry involves tissue fixation, histological sectioning, antibody incubations and confocal microscopy.

Project availability: Michaelmas and Lent Term

Other relevant themes: World class underpinning bioscience

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Improved Root Traits in Wheat

Supervisor: Dr. Emma Wallington, National Institute of Agricultural Botany

Project abstract: We would like to offer a 10 week project to study potential improved root traits in wheat, extending published work from other species. The aim is to increase wheat root length, and thus improve water use efficiency, to improve P, N & K uptake and therefore yield. We have transformed wheat with constructs to constitutively express wheat homologues of genes involved in these root traits, or to reduce their expression by RNAi-silencing. The 10 week DTP project will initially genotype and phenotype these materials, identify homozygous plants, then look at the effect on gene expression.

Learning outcomes and skills acquired: Laboratory skills will include molecular analysis of transformed wheat plants (PCR, Q-PCR and RT-PCR).

Project availability: Michaelmas and Lent Term

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Hormonal regulation of root mechanosensing

Supervisor: Dr. Julia Davies, Plant Sciences

Project abstract: As roots penetrate the soil they undergo mechanical stimulation that can effect changes in root system architecture. This is particularly relevant to the quest for roots better adapted to dry soil. In Arabidopsis, mechanical stimulation of roots causes transient elevation of cytosolic free calcium as a second messenger. This involves activation of plasma membrane mechanosensitive channels. It also  causes release of ATP to the apoplast where it can act as a cell growth regulator (1) Ultimately, there is  transcription of touch-responsive genes and alterations in root growth. Our preliminary findings show that strigolactones (2) can suppress mechanically induced elevation of free calcium and ATP release in Arabidopsis roots. This is a new role for this class of plant hormones. We have transformed strigolactone synthesis and sensing mutants to express aequorin or YC3.6 as indicators of cytosolic free calcium. We now need to establish whether these mutants have an aberrant calcium increase in response to mechanical stimulation. Additionally, we need to determine their level of ATP release, transcription of touch responsive genes and growth responses. Finally, we need  to establish which mechansosensitive channels are involved in strigolactone-sensitive ATP release.  A range of techniques will be used from imaging to qPCR.    (1)Science 343, 290 (2)Nature 455, 195

Learning outcomes and skills acquired: The student will gain an appreciation of signalling mechanisms, particularly thos involving calcium. Key skills will include luminometry, confocal microscopy, image analysis, and qPCR.

Project availability: Lent Term Only (January - March 2015)

Other relevant themes: World class underpinning bioscience

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Regulation of genes underpinning improved photosynthetic performance

Supervisor: Professor Julian Hibberd, Plant Sciences

Second supervisor: H Griffiths, Plant Sciences

Project abstract: C4 photosynthesis is a modified carbon fixation pathway that enhances growth. It involves the accumulation of several enzymes to high levels in either mesophyll or bundle sheath cells (Hibberd & Covshoff, 2010). Identifying mechanisms responsible for these patterns of expression would provide insight into the evolution of C4 photosynthesis, but also impact on strategies being taken to engineer the C4 pathway into crop species such as rice. Two genes that require high levels of expression for C4 photosynthesis in C. gynandra are those encoding NAD-malic enzyme (CgNAD-ME1) and a β-carbonic anhydrase (CgCA4). You will determine chromatin modification state across the both genes in M and BS cells and relate this to regions known to be sufficient to increase expression (Brown et al., 2010). The second aim of this project is to assess the importance of introns in regulating the expression levels of CgCA4. Introns have been shown to influence the expression levels of eukaryotic genes through a variety of mechanisms (Le Hir et al, 2003). CgCA4 evolved to be significantly shorter than its Arabidopsis counterpart through the truncation of intron sequences. To test whether these changes have functional significance in the expression of CgCA4, you will prepare reporter fusions of the Arabidopsis and C. gynandra genes with and without introns so that GUS accumulation can be quantified as outlined above.

Learning outcomes and skills acquired: This project will familiarise you with many essential skills in molecular biology, including the extraction of DNA and protein, polymerase chain reaction (PCR), protein quantification and cloning. The results will improve our understanding of the evolution of C4 photosynthesis and may have applications in the genetic engineering of novel C4 crop varieties.

Project availability: Michaelmas and Lent Term

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Characterisation of von Willebrand factor-binding protein in bovine and human isolates of MRSA.

Supervisor: Dr. Mark Holmes, Veterinary Medicine

Second supervisor: Dr. Ewan Harrison, Veterinary Medicine

Project abstract: Currently in Dr Mark Holmes’ group a large project is underway investigating the prevalence and diversity of Staphyloccocus aureus strains carrying a novel mecA homologue conferring methicillin resistance (MRSA) [1]. As part of this study a large number of strains isolated from both human and bovine infections have undergone whole genome sequencing. To further understand the evolutionary history of isolates we have begun investigating a number of genes that are known to be involved in the host-specificity of S. aureus. One particular gene is vWbp, which encodes von Willebrand factor-binding protein (vWbp). vWbp is a virulence factor that binds to pro-thrombin causing its activation and leading to the conversion of fibrinogen to fibrin causing the clotting of blood [2,3].  Along with other proteins vWbp promotes the formation of fibrin clumps, which probably help protect the invading S. aureus from phagocytosis [4]. Recently, it has been demonstrated that variants of vWbp have varying abilities to coagulate different animal plasmas and that this specificity correlates with the host the strains were isolated [4]. Thus the specificity of vWbp proteins can be used to infer the host specificity and evolutionary history of the S. aureus strains that harbour them. We have identified from whole genome sequencing that the two major clones of S. aureus that carry the novel-mecA homologue both have divergent vWbp, which are unique compared to other vWbp characterised to date. This project will aim to ascertain if these novel variants of vWbp can coagulate human and/or animal (bovine) plasmas. This will help provide us with evidence to identify if these strains are adapted to humans or cows, thus giving insight into the strains likely origins and direction of transmission (humans to cows or cows to humans). This project will initially focus on using data generated from whole genome sequencing to determine the diversity and phylogenetic  relationships of isolates possessing the vWbp gene. A panel of S. aureus strains will be tested against a panel of animal plasmas to identify if these strains can coagulate both human and animal plasmas. Subsequently attempts will be made to clone the genes from the   S. aureus host strains that have host specific vWbp variants and use recombinate DNA technology to transfer them into another S. aureus strain which has no coagulase activity in order to provide definitive evidence of function and host specificity. 

References

1. Garcia-Alvarez L, et al. (2011). Lancet Infect Dis 11: 595-603.

2. McAdow M, et al. (2012). J Innate Immun 4: 141-148.

3. Kroh HK, et al. (2009). Proc Natl Acad Sci U S A 106: 7786-7791.

4. Viana D, et al. (2010). Mol Microbiol 77: 1583-1594.

Learning outcomes and skills acquired: The project student will gain experience and skills in the following areas: (1) Searching and analysis of genetic data derived from whole genome sequencing (de novo assembly of Illumina fastq data, BLAST searching, SNP analysis etc.) (2) Conventional laboratory microbiology (culture and phenotypic characterisation of S. aureus isolates) (3) Use and refinement of plasma clotting assays (4) Use of molecular biology to manipulate S. aureus genomes (use of recombinate DNA technology to delete and transfer genes between S. aureus strains). (5) Experimental design, appropriate statistical analysis and interpretation of results.

Project availability: Michaelmas and Lent Term

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Totipotent stem cells: from mouse to human

Supervisor: Dr. Pentao Liu, The Wellcome Trust Sanger Institute

Project abstract: Mouse embryonic stem (ES) cells from the blastocyst can differentiate to all three somatic germ layers, and when returned to the blastocyst environment, they are able to efficiently colonise all germ layers of the embryo. In the last couple of decades, mouse ES cells have revolutionised studies of developmental biology, genetics, and human disease mechanisms in the mouse. Human ES cells hold great promise in regenerative medicine and cell-based therapies. However, ES cells are not available from most other mammalian species including agriculturally important species such as pig and cow. This is partly due to species differences but it also reflects our lack of understanding of the fundamental aspects of genetics and signaling in preimplantation embryos in mammals. The zygote or blastomeres of 2-cell to 8-cell mouse embryo have the capacity to differentiate to both embryonic and placenta tissues and are considered totipotent. In a soon to be published study, we have successfully established, for the first time, homogenous culture of cells with expanded developmental potential (totipotent) that are isolated directly from mouse preimplantation embryos. Functional and molecular analyses map these cells to being similar to blastomeres of 4-cell to 8-cell embryos. These exciting results suggest that similar cells of other mammalian species may also be captured and maintained in long-term culture. In this rotation project, you will participate in further characterization of totipotent stem cells of mouse and also of other mammalian species. The training will include stem cell biology, bioinformatics (single-cell RNA-seq, ChIP-seq) and other lab skills.

Learning outcomes and skills acquired: Stem cell knowledge in particular on totipotent stem cells, basic experimental and bioinformatics skills. Since we are the first to establish homogenous totipotent cell cultures, you will have ample opportunities to explore and expand your studies.

Project availability: Michaelmas and Lent Term

Other relevant themes: Basic bioscience underpinning health & World class underpinning bioscience

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Towards an invertebrate model of transmissible mammalian prion disease

Supervisor: Dr. Raymond Bujdoso, Veterinary Medicine

Second supervisor: Dr.. Alana Thackray, Veterinary Medicine

Project abstract: Prion diseases are infectious fatal neurodegenerative conditions of humans and various animal species. These conditions include scrapie in sheep, bovine spongiform encephalopathy (BSE) in cattle and Creutzfeldt-Jakob disease (CJD) in humans. Many fundamental issues relating to the molecular pathogenesis of prion diseases remain poorly defined. These include: an understanding of the molecular nature of the infectious prion agent; the mechanism of prion propagation and the mechanism of PrP-mediated -neurotoxicity.     We have investigated Drosophila melanogaster as a new animal model of ovine scrapie in order to provide a more tractable model of transmissible mammalian prion disease. This PhD project will contribute to the establishment of the fruitfly as an invertebrate animal model of transmissible mammalian prion disease.     The aim of this proposal is to test the hypothesis that PrP transgenic Drosophila are a suitable invertebrate host to measure mammalian prion infectivity.

The specific objectives are:

1. Determine the sensitivity of ovine PrP transgenic Drosophila to ovine prion infectivity  

2. Use ovine PrP transgenic Drosophila to measure prion infectivity in brain, lymphoid tissue and blood from scrapie-infected sheep in order to replace the use of mammals in the current bioassays  

3. Strain type ovine prions in ovine PrP transgenic Drosophila   

References 

1. Thackray, A. M., et al.  2012. Prion-induced toxicity in PrP transgenic Drosophila.  Exp Mol Pathol 92(2): 194-201. 

2. Thackray, A. M., et al. 2012. Ovine PrP transgenic Drosophila show reduced locomotor activity and decreased survival. Biochem J 444(3): 487-495.

Learning outcomes and skills acquired: This project will develop skills in: molecular biology, protein biology, transgenesis in Drosophila, prion biology, animal models of neurodegeneration, experimental design and methodology, data handling and analysis, transferable skills such as team work, data presentation and communication.

Project availability: Michaelmas and Lent Term

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A compatibility factor in plant colonisation by symbionts and pathogens: From characterisation to deployment

Supervisor: Dr. Sebastian Schornack, Sainsbury Laboratory

Second supervisor: Dr. Thomas Rey, Sainsbury Laboratory

Project abstract: The majority of all land plants is able to engage in symbiosis with root colonising beneficial mycorrhizal fungi. This interaction is supported by a conserved plant genetic network required to suppress immunity and trigger development related mechanisms associated with microbial accommodation. Whether such processes can also be utilized by filamentous pathogens to colonize plant host tissues is still unresolved, notably because of the lack of pathosystems in symbiotic plants.    We established interaction assays between the oomycete Phytophthora palmivora and the legume Medicago truncatula. Screenings of symbiosis defective mutants for resistance to the pathogen led to identification of the api mutant conferring incompatibility to the disease. Dissection of the colonisation process showed that API is required for successful penetration events at the  rhizodermis.    The aims of the proposed internship will be to 1) characterize how API mediates the penetration by the pathogen and to 2) investigate the transferability of the resistance related to API knock out in leaves and roots of crop plants.    You will design constructs enabling confocal imaging of cellular responses to the colonisation by P. palmivora in leaves and roots. Transgenic roots will also be challenged with mycorrhizal fungus to compare API dependent cellular responses in these contrasted interactions.    You will knock-out tobacco API homologs by CRISPR/Cas9 genome editing. Transgenic plants will be challenged with Phytophthora in leaves and roots and also tested for mycorrhization. Alternatively you will generate a dominant negative version of API that will be overexpressed in plants to mimic phenotypes obtained from api mutant.

Learning outcomes and skills acquired: During your research you will get the opportunity to acquire and employ several plant-microbe research techniques, including: Agrobacterium-mediated transformation of tissues and plants, fluorescence (confocal) microscopy, scanning electron microscopy, atomic force microscopy, maintenance of microbial pathogens and infection assays, transformation and regeneration of Phytophthora. You will also gain mycorrhizal, plant and fungal biology knowledge. Furthermore you will use state-of-the-art genome editing technology such as TAL effectors or CRISPR/Cas9 to genetically engineer plants with modified interaction potential.    As a candidate you should be familiar with standard molecular biology techniques such as PCR, cloning and E. coli work.

Project availability: Michaelmas and Lent Term

Other relevant themes: World class underpinning bioscience

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Interference between oomycete effectors and formation of mycorrhiza

Supervisor: Dr. Sebastian Schornack, Sainsbury Laboratory

Second supervisor: Dr. Edouard Evangelisti, Sainsbury Laboratory

Project abstract: Plants interact with microorganisms, especially in the rhizosphere where root exudates attract many microbes. Some of these microbes engage in mutually beneficial interactions, whereas plant pathogens invade root tissues and cause diseases. Both types of interactions rely on molecular exchanges between the partners. Pathogenic microbes have evolved small secreted proteins termed effectors that promote infection by interfering with host processes, including immune responses. Beneficial fungi also secrete effectors: the basidiomycete Laccaria bicolor secretes the effector MiSSP7 during interaction with poplar to favour mycorrhiza formation (Plett et al, 2014).   In the lab we are focusing on the plant-pathogenic oomycete Phytophthora palmivora which infects more than 200 plant species, including important crops such as oil palm and coconut. P. palmivora also infects tobacco (Nicotiana benthamiana), a species which is able to develop mycorrhiza with the fungus Rhizophagus irregularis.    During your internship you will use the N. benthamiana–R. irregularis system to investigate the effect of P. palmivora effectors on the establishment of mycorrhiza. This work will identify plant processes that may be targeted differentially by effectors from beneficial and detrimental microorganisms. You will also be involved in the in silico identification of secreted Rhizophagus effectors and initiate their functional analysis.                As a candidate you should be familiar with standard molecular biology techniques such as PCR, cloning and E. coli work.

Learning outcomes and skills acquired: During your research you will acquire and employ plant-microbe research techniques including: Tobacco-Rattle-Virus (TRV)-mediated expression, Agrobacterium-mediated transformation, fluorescence microscopy and bioinformatics tools. You will also gain plant and fungal biology knowledge.

Project availability: Michaelmas and Lent Term

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Determining targets of anti-fecundity immunity in Schistosoma bovis infection

Supervisor: Dr. Shona Wilson, Pathology

Project abstract: Schistosoma bovis is an infection of ruminants in Africa, the middle East and the Mediterranean.  In some foci in sub-Saharan Africa, the prevalence and intensity of infection in cattle can be very high.  Infection causes poor health, resulting in a reduction in productivity, hindered transport to market, as this is often conducted on foot, and can result in early slaughter, particularly in times of adverse conditions.   S. bovis belongs to a clade of the Schistosoma genus that also contains S. haematobium, the schistosome arguably of the greatest public health importance in humans.  The two infections show similar characteristics, including a gradual development of immunity, so the young carry the greatest infection burden (assessed by egg output). For both infections, this immunity is partly attributable to reduced egg production by the paired adult worms. Due to the close relationship between the two species, recently shown to form hybrids, the targets of anti-fecundity immunity in the two infections are likely to have a high degree of identity.  In this project a plasma set from Malian children infected with S. haematobium, for whom egg to worm ratios are known, will be used to investigate targets of anti-fecundity immunity by 2D Western blot analysis.  S. bovis orthologues of targets will be identified from the newly sequenced S. bovis genome.  Long-term aims are to clone and express these S. bovis orthologues and to screen Ab responses amongst cattle raised in Mwanza Region, Tanzania.

Learning outcomes and skills acquired: The project will have a case-control design. The student will define cases and controls from egg output to worm burden ratios of 62 Malian adults, and select the cohort, using propensity score modeling to match on age, sex, tribal group and village of residence, from 131 Malian children, participants in a wider cohort.    The running of 2D gels/mass spectometry will be outsourced, but the student will conduct the Western blots.  As this will involves handling of human plasma samples HepB vaccination is required.      The student will identify S. haematobium targets from the published genome and blast for S. bovis orthologues.

Project availability: Lent Term Only (January - March 2015)

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The effect of Ppd on stress tolerance at germination and establishment in wheat

Supervisor: Dr. A Bentley, National Institute of Agricultural Botany

Second supervisor: Dr. I Mackay, National Institute of Agricultural Botany

Project abstract: Ppd is a locus in cereals characterised primarily by its major effect on flowering time and height. However, it is also known to influence frost tolerance and winter kill. This project will use a set of near isogenic lines in wheat to assess if an effect of Ppd-D1 can be detected on germination and coleoptile emergence rates in laboratory tests. The tests will then be extended to impose water logging, drought and freezing tests on seeds and seedlings. The work will be extended to survey variation across parents of mapping population and representatives of adapted UK germplasm prior to initiating linkage and association mapping experiments.

Project availability: Michaelmas and Lent Term

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Ergot susceptibility in hexaploid wheat: analysis of transcriptome timecourse

Supervisor: Dr. Anna Gordon, National Institute of Agricultural Botany

Project abstract: The Ergot fungus infects wheat via its’ floral tissue and within days colonises the whole flower to become a mycotoxin-rich fungal mass called an ergot. There is no known full resistance to this fungus and as such we have completed a detailed transcriptome study covering the first 7 days in a susceptible interaction. We are aiming to understand how the fungus invades and what the key moments are that determine the success of infection, and to understand what the plant resistance responses are. The initial analysis of the transcriptome data suggests that there are a large set of Auxin responsive genes that are induced by 3 days after inoculation, implying that perturbation of the plants auxin homeostasis could be the target of the pathogen in order to facilitate a successful colonisation.

Learning outcomes and skills acquired: This 10-week DTP project will use a set of bioinformatic tools to analyse our vast dataset to answer relevant biological questions about the progression of disease. There is scope for carrying out RT-PCR to confirm changes in gene expression patterns. The student will also have a chance to clone a fragment of an auxin responsive gene into an RNAi entry vector destined for plant transformation and to genotype GM wheat plants already in production.

Project availability: Michaelmas and Lent Term

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Capturing Photosynthetic Efficiency Traits from Tetraploid Wheat

Supervisor: Dr. Fiona Leigh, National Institute of Agricultural Botany

Project abstract: Ancestral wheat species harbour novel genetic variation that we hope to capture in wheat improvement efforts. At NIAB we are intercrossing a range of tetraploid wheats (Triticum durum, T. dicoccum, and T. dicoccoides) with modern hexaploid bread wheat (T. aestivum), through resynthesis and direct crossing, as part of WISP, the BBSRC public-good wheat pre-breeding program (http://www.wheatisp.org).   Diversity in photosynthetic and water use efficiency (WUE) traits was observed in our collection of tetraploid wheats, extending above and below the values obtained for elite bread wheats.  This PhD will interrogate the tetraploid gene pool with the ambition of improving the photosynthetic capacity of bread wheat in future varieties.

Learning outcomes and skills acquired: The research will involve: a) development of novel mapping populations from reciprocal crosses between parents which contrast for key traits. b) phenotyping of these populations with gas exchange techniques to establish the heritability of photosynthesis and WUE traits. c) identification of quantitative trait loci (QTL) using high density molecular markers and bioinformatic tools to begin trait dissection. d) investigation of the influence of influence of the cytoplasm in the expression of photosynthesis / WUE phenotypes

Project availability: Michaelmas and Lent Term

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Deploying next-generation biological resources and dense SNP chips for the genetic dissection of brown rust resistance in wheat

Supervisor: Dr. I Mackay, National Institute of Agricultural Botany

Second supervisor: Dr. James Cockram, National Institute of Agricultural Botany

Project abstract: This project will deploy the NIAB 8 founder elite wheat MAGIC population and the dense 90k genotype dataset to investigate the genetic basis of brown rust resistance, a major wheat pathogen in the UK and worldwide. Brown rust is caused by the fungal pathogen Puccina recondita, and can reduce yields by up to 50%. As leaders of the UK Cereal Pathogen Virulence Survey, NIAB possesses the national brown rust isolate collection. Using the expertise and resources available, this project will undertake replicated phenotypic tests for seedling resistance to specific brown rust isolates under controlled environment conditions. Subsequently, the phenotypic data generated will be combined with the existing 90k SNP data to identify genetic components of brown rust resistance. The genomic context of these markers will be investigated using emerging wheat physical and sequence resources and comparative genomic approaches, and breeder-friendly genetic markers generated, as time permits. Previous screens for yellow rust resistance in the MAGIC population have identified highly significantly associated genetic markers, and interestingly, transgressive segregation for resistance. This work package forms part of a wider objective of utilising wheat MAGIC for the efficient application of genomics-assisted plant breeding for sustainable wheat production, targeting (but not limited to) disease resistance and yield components within a unified genetic mapping platform.

Project availability: Michaelmas and Lent Term

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PhD opportunities in Statistical and Quantitative Genetics

Supervisor: Dr. I Mackay, National Institute of Agricultural Botany

Project abstract: Covering the application of methods from statistical, population and quantitative genetics to improve the efficiency of plant breeding, NIAB can offer projects ranging from the development of novel methods and algorithms for trait mapping to more practical lab & field based experimentation including: association mapping in wheat and barley, development and application of methods for genomic selection in autogamous species, processes for improving the speed and efficiency of inbreeding in crop breeding, the Multiparent Advanced Generation Intercross (MAGIC), the design of breeding programmes.

Project availability: Michaelmas and Lent Term

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The use of historical trials data to track genetic and environmental trends in UK crops

Supervisor: Dr. I Mackay, National Institute of Agricultural Botany

Project abstract: Published work (Mackay et al.  2010) in which historical data from variety trials was reanalysed, demonstrated that from 1982 to 2007 the most important component of yield increase in the major UK arable crops was genetic rather than environmental (arising from either climate change or changes in crop agronomy). These analyses can be updated with the addition of a further seven years’ data. In addition to historical trials data, there are also several datasets available in which old varieties have been compared with new varieties in the same trial series. These trials can be analysed separately and also combined with the historical trials series to assess bias in the reanalysis of historical data and the extent to which varieties have been bred not only for increased yield but also for adaptation to the prevalent environment. The original analyses indicated that annual weather records could be used to identify drivers of variety instability over years. There are more data available on variation between trial locations within years and the extension of the analyses to incorporate these should give a more accurate assessment of the causes of variety instability.

Project availability: Michaelmas and Lent Term

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PhD opportunities in Statistical and Quantitative Genetics

Supervisor: Dr. I Mackay, National Institute of Agricultural Botany

Project abstract: Covering the application of methods from statistical, population and quantitative genetics to improve the efficiency of plant breeding, NIAB can offer projects ranging from the development of novel methods and algorithms for trait mapping to more practical lab & field based experimentation including: association mapping in wheat and barley, development and application of methods for genomic selection in autogamous species, processes for improving the speed and efficiency of inbreeding in crop breeding. the Multiparent Advanced Generation Intercross (MAGIC), the design of breeding programmes

Project availability: Michaelmas and Lent Term

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A comparison of genomic selection and marker assisted selection in a bi-parental cross

Supervisor: Dr. K Gardner, National Institute of Agricultural Botany

Second supervisor: Dr. I Mackay, National Institute of Agricultural Botany

Project abstract: Genomic selection is a method of breeding in which large numbers of markers are used to predict traits without recourse to identifying individual QTL through linkage or association mapping. In marker-assisted selection, linkage and association mapping must first identify individual markers with statistically significant effects on target traits. Selection then takes place on a score based on these markers alone. Using cross-validation approaches this project will compare genomic selection and marker assisted selection in bi-parental crosses and in association mapping panels of winter wheat.

Project availability: Michaelmas and Lent Term

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Comparison of genetic and pedigree based relationships in the UK winter wheat pedigree

Supervisor: Dr. K Gardner, National Institute of Agricultural Botany

Second supervisor: Dr. I Mackay, National Institute of Agricultural Botany

Project abstract: We have catalogued the common pedigree of contemporary UK winter wheat lines stretching back over 50 years. High density marker data are available on many of the contemporary and ancestral lines. This project will compare relationships among lines computed from markers and from the pedigree. Results will be used to test the accuracy of the pedigree, to compare the use of pedigree and marker based relationship matrices in association mapping and to search for regions of the genome from ancestral lines which appear overrepresented in their descendents.

Project availability: Michaelmas and Lent Term

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Developing genetic markers for wheat domestication traits

Supervisor: Dr. Phil Howell, National Institute of Agricultural Botany

Project abstract: NIAB is providing the ‘synthetics pillar’ of WISP, a BBSRC public-good wheat pre-breeding programme (http://www.wheatisp.org), which involves developing two parallel streams of wheat pre-breeding material.  The first is based upon resynthesis (a recreation of the rare hybridisation event that led to the emergence of hexaploid wheat 10 000 years ago, see http://www.niab.com/uploads/files/NIAB_Synthetic_Hexaploid_Wheat.pdf), and the second involves direct crossing between tetraploids and hexaploids. Through this work, we are introducing diversity from a range of wild and cultivated diploid (Aegilops tauschii, DD genome) and tetraploid (Triticum durum, T. dicoccoides, T. dicoccum, all AABB) species into elite hexaploid wheat varieties (T. aestivum, AABBDD). Many of the lines we produce will display many undesirable “weedy” characteristics, such as shattering seed heads or adhering glumes. Shattering is caused by the homoeologous Br loci (Watanabe et al., 2006. J Appl Genet 47: 93-98), whilst tenacious glumes are controlled by the Sog and Tg loci (Sood et al., 2009. Theor Appl Genet 119: 341-351). The most important domestication gene, the Q locus, has been cloned and appears to belong to the AP2 family of transcription factors (Simons et al., 2006. Genetics 172: 547-555). Genetic markers for these characters will greatly aid the selection of improved pre-breeding material and help widen the pool of diversity available to commercial wheat breeders, ultimately leading to improvements in yield stability, pest and disease resistance, and the tolerance of abiotic stresses. Diploid, tetraploid and hexaploid materials from our WISP programme can be characterised for threshability and then screened with published markers linked to these major domestication loci, together with additional SNP-based markers developed by WISP genotyping partners.  Existing segregating populations will be used to develop co-dominant markers suitable for high-throughput genotyping. This is likely to require a bio-informatics approach exploiting synteny with rice, maize and brachypodium. Once very close linkage is established, it should be possible to develop near-isogenic lines for these loci in order to better understand the underlying mechanism of each character and how they interact. As well as making the pool of non-domesticated wheat relatives more accessible to breeders, this may give some new indications on how to optimise the balance between desirable threshability and undesirable grain shedding.

Project availability: Michaelmas and Lent Term

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Grain cadmium levels in tetraploid and hexaploid wheat

Supervisor: Dr. Phil Howell, National Institute of Agricultural Botany

Project abstract: The NIAB pre-breeding group are intercrossing tetraploid wheats (Triticum durum, T. dicoccum and T. dicoccoides) with hexaploid bread wheat (T. aestivum), as part of WISP, the BBSRC public-good wheat pre-breeding programme (http://www.wheatisp.org). Grain samples of tetraploid wheat have consistently shown higher levels of the toxic heavy metal cadmium (Cd) than hexaploid wheat, and low-Cd accumulation is now a trait of major importance to durum wheat breeders. However, a significant proportion of recent UK bread wheat varieties have tetraploid parentage in their pedigrees, largely coming from T. dicoccoides, which has unknown Cd status. In addition, our WISP pre-breeding work may be inadvertently transferring high-Cd alleles from tetraploid wheat into low-Cd hexaploid wheat. Recent work has identified a locus (Cdu1) which explained 82% of the phenotypic variation in Cd accumulation in a segregating durum wheat population, and which has been mapped to a 0.7cM interval on chromosome 5BL (Wiebe et al., 2010. Theor Appl Genet 121:1047-1058). Flanking markers can be used to screen a large range of tetraploid and hexaploid accessions from our WISP crossing work. Wholegrain flour samples, originating from field trials of these accessions, can be prepared for ICP-MS analysis (in conjunction with the Department of Earth Sciences) to determine Cd levels. This screening approach will allow a detailed genetic analysis of cadmium uptake in our WISP material. The durum Cdu1 interval corresponds to a relatively small region (approx. 285 kbp) in both rice and brachypodium (Wiebe et al., 2010), so the development of diagnostic genetic markers suitable for high-throughput genotyping in wheat should be possible. These can then be used for routine selection against high-Cd alleles in segregating WISP populations, and transferred to commercial breeding programmes as necessary. Cadmium accumulation may also be a suitable phenotype for exploring changes in gene expression following polyploidisation. For example, would a hexaploid derived from a high-Cd tetraploid through re-synthesis necessarily accumulate the same Cd levels as its tetraploid progenitor? What is the role of the (presumably non-functional) Cdu1 homoeologues on the A- and D-genomes?

Project availability: Michaelmas and Lent Term

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The Chocolate spot toolbox

Supervisor: Dr Anne Webb, National Institute of Agricultural Botany

Project abstract: Chocolate spot (Botrytis fabae) is an important disease of the emerging legume crop faba bean (Vicia faba). It causes severe crop losses and there are currently no fully resistant commercial cultivars or resistance genes available. NIAB has assembled a large collection (>900 accessions and inbred lines) of V. faba, including lines partially resistant to chocolate spot, as well as >200 isolates of B. fabae from around the UK. This 10-week DTP rotation project provides an exciting opportunity to initiate parallel research into resistance sources and pathogen dynamics, providing the basis for future breeding towards resistance.

This will be achieved through:  

1. Identification of sources of resistance to chocolate spot. Faba bean lines will be inoculated with B. fabae in a controlled environment to determine the range of natural exploitable resistance available. 

2. Exploration of the genetic diversity within NIAB’s collection of B. fabae isolates. The sequenced genome of the extensively characterised and closely related pathogen B. cinerea will be used to develop molecular markers for validation in B. fabae.

Learning outcomes and skills acquired: Skills essential to plant pathology: identification, cultivation and storage of plant pathogenic fungi in sterile and non-sterile environments, set-up, running and evaluation of inoculated variety trials and the interpretation of results.   Biotechnology-related skills: use of bioinformatics tools to identify molecular markers, testing, set up and running of marker assays, practice of essential molecular procedures, e.g. DNA extraction and PCR.  Generic research skills: set up of experiments, analysis and interpretation of results, documentation of methods and data.

Project availability: Michaelmas and Lent Term

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Developing new plant-breeding tools for the long-chain, omega-3 oil crop Buglossoidies arvensis

Supervisor: Dr Thomas Wood, National Institute of Agricultural Botany

Second supervisor: Dr Steven Bentley, National Institute of Agricultural Botany

Project abstract: The inclusion of fish oils containing long chain omega-3 poly-unsaturated fatty acids (PUFAs) has been shown to have been associated with reduced rates of cardio-vascular disease. Buglossoides arvensis (Corn Gromwell), a previous weed of cereal crops, is a new spring crop for the U.K. which provides a sustainable, non-GM source of stearidonic acid (SDA). SDA is (one) precursor in the biosynthesis of the LC-PUFAs eicosatetraenoic acid (ETA 20:4 n-3), eicosapentaenoic acid (EPA, 20:5 n-3) and docosahexaenoic acid (DHA, 22:6 n-3), commonly found in fish oils (Ruiz-Lopez et al., 2014 Plant J 77: 198-208). NIAB, in collaboration with Tech Crops International (TCI), is developing* new B. arvensis varieties for commercial production of SDA. Although a relatively diverse collection of B. arvensis germplasm is available (to the breeder), current genetic resources are limited and breeding must rely on classical methods to develop new, improved varieties. This project aims to develop a series of inter-simple sequence repeat (ISSR) markers (Korpelainen et al., 2007 BioTechniques 42: 479-486) to screen the B. arvensis collection held at NIAB. Using a series of generic ISSR primers, PCR products will be directly sequenced to obtain information adjacent to SSR sites to develop new molecular markers. New markers will then be used to establish any phylogenetic relationships and possible population structure between accessions. The development of new molecular tools will facilitate the selection of improved B. arvensis lines and also aid in the identifying genes controlling keys traits such as oil yield or profile. The DTP placement will provide the candidate with the opportunity to experience practical development of breeding tools with potential for direct application to a new U.K. crop.

Project availability: Michaelmas and Lent Term

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The genetic correlates of dietary wariness in poultry

Supervisor: Dr Hannah Rowland, Zoology

Project abstract: This project addresses a fundamental question about managed-animal nutrition: what are the genes underlying dietary wariness? In a commercial context, broiler chickens are given several changes in diet over the course of their development. These changes are associated with a refusal by some individuals to continue eating. Termed dietary wariness (DW), this lasting refusal to accept novel food into the diet causes a considerable reduction in animal welfare, growth rate and productivity (Cooper 1971; Murphy and Duncan 1977; Poole 1999). DC has been demonstrated in eight species of birds (Marples et al., 2005) and in five species of fish (Thomas et al., 2010;  Richards et al., 2011; Richards et al. 2014) and has been shown to be heritable (Marples and Brakefield 1995). You will measure the behaviour of chickens and classify their level of dietary wariness. You will then use a candidate gene approach to assess the association between specific alleles of the family of bitter taste receptor genes that have been implicated in the expression of DW. Understanding the mechanism underlying responses to dietary change could be used to improve the welfare and productivity of captive poultry. A greater understanding of DW is also important for conservation because changes in climate and human activities such as farming practices are altering the foods available to many species  and changing their geographical distribution

Learning outcomes and skills acquired: The student will conduct research that will inform strategies for improving the conditions and management of farmed, laboratory, and other managed animals. They will gain skills in experimental design, behaviour recording, DNA extraction and PCR. The student will also have the opportunity to contact multidisciplinary collaborators.

Project availability: Lent Term Only (January - March 2015)

Other relevant themes: Basic bioscience underpinning health

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Diffusion of microorganisms in soil

Supervisor: Dr S Taraskin, Chemistry

Project abstract: Understanding how cooperative and non-linear phenomena influence the movement and spatial distribution of biota in    natural habitats such as soil is an important and fascinating question relevant to several disciplines.   The soil pore space is a highly heterogeneous habitat hosting a stunning wealth of biological activity (e.g. of bacteria,   fungi or nematodes)  that plays an essential role in many processes including plant growth, climate change, or soil-borne epidemics.    The aim of this project is to study theoretically (both numerically and analytically) spread of microorganisms through soil.   The simplest mathematical model which can describe the spread is the diffusion model.   In this model, an organism can stochastically move between pores in soil.   The diffusional jumps can be biased in a certain direction due to gravity or   presence of nutrients.   The fractal topology of soil brings additional interesting features to diffusion.   Real soil samples with and without tillage treatment will be used to constract a network model of soil [1].   Soil networks consist of a set of nodes and edges whose layout captures the  topology of the soil pore space where biological activity takes place.  The network representation is achieved by associating the branching points of the soil pore space  with the nodes and the pore-space channels between branching points with the   network edges.   The diffusion model defined on such a network can be solved approximately analytically   ignoring the correlations in transmissibilities and exactly numerically   by means of the Monte Carlo simulations.     The main outcome of the project will be an estimate of typical time for organisms of different size to invade soil up to certain depth.   Mathematically, the power-low dependence of the mean-squared displacement on time is the main objective of the analysis.   The knowledge of such dependence would allow the effect of the strength of tillage to be studied and used for prediction   and control of invasion in soil.    

Perez-Reche F.J., Taraskin S.N., Otten W., Viana M.P., Costa L.D.F., Gilligan C.A., Phys. Rev. Lett. 109, 098102 (2012)

Learning outcomes and skills acquired: The student will learn:   (i) how to formulate diffusion models for studying microbial invasion in   complex networks representing soil,   (ii) how to solve analytically (using mean-field) simple diffusion models,   (iii) how to use numerical techniques to simulate diffusion and similar spreading phenomena in lattice models   and complex networks,  (iv) how to construct complex networks and analyse their topological properties,   (v) how to estimate the diffusion coefficient for microbial invasion and relate it   to structural and topological characteristics of the underlying network.

Project availability: Michaelmas and Lent Term

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Does the enhanced resistance of wheat against rust fungi impact on interaction with beneficial fungi?

Supervisor: Dr Uta Paszkowski, Plant Sciences

Second supervisor: Emma Wallington National Institute of Agricultural Botany

Project abstract: The development of plant diseases requires susceptible plants and adapted invaders, hence compatible organisms. In contrast, incompatibility results in resistance. The disease resistance genes Lr34 (1) and Yr36 (2) from bread wheat confer durable protection against a variety of devastating fungal rust pathogens, and have therefore been highly relevant for wheat breeding since the green revolution. Both genes have been cloned and refer to an ABC-transporter of the pleiotropic drug resistance subfamily and a kinase with a START lipid-binding domain. Rust fungi are obligate biotrophs that infect above ground plant parts and require the living host to complete their life cycle. Also arbuscular mycorrhizal (AM) fungi are obligate biotrophs; however the fungus proliferates in the root of the host and the outcome of the interaction is mutualistic. It has been shown that there are plant compatibility modules shared between AM fungi and taxonomically distant organisms (3). The effect of Lr34 and Yr36 on root interaction with beneficial fungi is currently unknown. It is the aim of this project to determine the effect of Lr34- and Yr36-mediated resistance on the development of arbuscular mycorrhizal symbiosis of wheat with Rhizophagus irregularis. You will learn a variety of methods for the characterization of the material; these will include performing mycorrhizal inoculation assays as well as histology, microscopy, plant molecular biology and genetics. You will be co-supervised by Emma Wallington at NIAB and by Uta Paszkowski at the Plant Science department, which will expose you to an “applied” and a “fundamental” research environment.

References:

1. S. G. Krattinger et al., A putative ABC transporter confers durable resistance to multiple fungal pathogens in wheat. Science 323, 1360 (Mar 6, 2009).

2. D. Fu et al., A kinase-START gene confers temperature-dependent resistance to wheat stripe rust. Science 323, 1357 (Mar 6, 2009).

3. E. Wang et al., A Common Signaling Process that Promotes Mycorrhizal and Oomycete Colonization of Plants. Curr Biol 22, 2242 (Dec 4, 2012).

Project availability: Michaelmas and Lent Term

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NO TITLE

Supervisor: Dr Uta Paszkowski, Plant Sciences

Second supervisor: Emma Wallington National Institute of Agricultural Botany

Project abstract: It has been widely documented that, under low input conditions, plants living in association with arbuscular mycorrhizal (AM) fungi grow bigger than their mock inoculated counterparts (1). This positive responsiveness can be attributed to the fungal mycelium enlarging the root surface area for nutrient acquisition. Responsiveness to AM colonization is defined as the difference in performance between colonized and non-colonized plants at a given level of nutrient availability (2). High responsiveness is an attractive agricultural trait, however, variable among plant species or even cultivars. To date, genetic determinants underpinning this phenotypic variation are elusive. Genetically diverse populations provide a powerful material for allele mining associated with traits of interest. Using the diverse eight founders of the wheat MAGIC (Multiparent Advanced Generation Inter-Cross, Mackay et al., submitted) population you will exploit this novel resource to genetically dissect variation in AM fungal colonization levels and mycorrhizal responsiveness. For this purpose you will inoculate the MAGIC founder panel with a defined number of spores from Rhizophagus irregularis to microscopically and molecularly determine fungal colonization. In addition, plant biomass production as a function of colonization will inform about the degree of responsiveness of eight weeks old plants, at a developmental stage prior flowering.

References:

1. S. E. Smith, D. J. Read, Mycorrhizal symbiosis.  (Academic Press, London, 1997).

2. D. P. Janos, Plant responsiveness to mycorrhizas differs from dependence upon mycorrhizas. Mycorrhiza 17, 75 (2007).

Learning outcomes and skills acquired: You will acquire a wide variety of expertise in crop genetics, plant molecular biology as well as in histology and microscopy. You will be co-supervised by Emma Wallington at NIAB and by Uta Paszkowski at the Plant Science department, which will expose you to an “applied” and a “fundamental” research environment.

Project availability: Michaelmas and Lent Term

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Visualizing endosymbiosis between rice and beneficial fungi

Supervisor: Dr Uta Paszkowski, Plant Sciences

Project abstract: Within the inner root layers of most land plants hyphae of ancient arbuscular mycorrhizal fungi (AMF) differentiate into tree-shaped feeding structures, called arbuscules (1). Monumental cellular re-differentiation in the inner cortex results in the de novo synthesis of a host-derived membrane that surrounds the arbuscule, called the peri-arbuscular membrane (PAM). This functional symbiosome interface governs the exchange of nutrients and signals between the two symbiotic partners.  Evidence that the PAM is distinct from the plant plasma membrane (PM) was obtained via Confocal Laser Scanning Microscopy (CLSM) using PM- and PAM-specific marker proteins fused to GFP and other fluorophores (2,3). However, cellular processes that underlie membrane dynamics during arbuscule development, maturation and senescence in the inner root cortex remain unclear and will be addressed in this study.  To this end, fluorescent marker lines are available in our laboratory to study subcellular compartments/organelles including PAM during arbuscule development.  Moreover, rice lines carrying functional GFP-tagged versions of the PAM-specific phosphate transporter OsPT11 (4) have been crossed with mRFP-tagged marker lines to enable co-localization.  You will use imaging and molecular tools to characterize progeny derived from crosses between various fluorescent subcellular marker lines. Co-localization of fluorescent marker proteins during arbuscule development will be done using CLSM. The data you will produce will provide valuable insights into the putative subcellular localization of PAM-specific proteins during arbuscule development. Overall, this project will equip you with key techniques in plant molecular biology and genetics, advanced CLSM imaging and 3D-image data handling.

References:

1. Parniske 2000 Curr Opin Pl Biol, 3, 320-328.

2. Pumplin and Harrison, 2009, Plant Phys, 151, 809-819.

3. Kobae and Hata, 2010, Plant Cell Physiol, 51 (3), 341-353.

4. Paszkowski et al, 2002, Proc Nat Acad Sci, 99 (22), 13324-13329

Project availability: Michaelmas and Lent Term

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Inhospitable: the importance of listening in plant-fungus conversations

Supervisor: Dr Uta Paszkowski, Plant Sciences

Project abstract: Plants can improve their provision of nutrients and water by establishing symbiosis with Arbuscular Mycorrhizal (AM) fungi, a fascinating phenomenon that still has to be fully understood. All the information available to date indicates that the very early stages of this interaction are critical for its successful outcome. Plant-fungus communication begins before physical contact and a suite of mobile signals from both the plant and the fungus have been implicated in the process.  The recent identification and initial characterization in our lab of the inhospitable (iho) rice mutant which is non responsive to the fungus in the first steps of the interaction, provides a unique tool for performing a highly focused comparison to gain a deeper insight in the earliest signalling events of AM symbiosis establishment.

Learning outcomes and skills acquired: During your rotation period in the lab you will be involved in defining the mechanisms of activation of the IHO signalling node. In particular you will be looking at protein and transcriptional dynamics in response to symbiotic stimuli. You will be performing the practical experimental work in collaboration with two experienced Postdoctoral fellows in the lab. This project will leave you with a solid training in molecular biology techniques of universal applicability such as molecular cloning, quantitative RT-PCR and yeast two hybrid assay. Furthermore, the participation to the regular scientific discussions in the lab will help you develop a deep understanding of the fundamental biological concepts underlying plant microbe interactions.

Project availability: Michaelmas and Lent Term

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Regulation of the pyrenoid-based carbon concentrating mechanism in Chlamydomonas

Supervisor: Professor Howard Griffiths, Plant Sciences

Project abstract: Many algae possess an inducible CO2 concentrating mechanism (CCM) to enhance the delivery of inorganic carbon (Ci) to Rubisco, thereby improving the operating efficiency of (arguably) the world's most fundamental protein by suppressing wasteful oxygenase activity. The algal CCM consists of three primary components: (i) various inorganic carbon membrane transporters, on both plasma- and chloroplast membranes; (ii) carbonic anhydrases (CA) in strategic locations, needed to catalyse the inter-conversion of CO2 and bicarbonate, either to maximize transport, storage or recapture of Ci supply; (iii) a pyrenoid, a chloroplast micro-compartment, within which Rubisco aggregates, which limits the back-diffusion (leakage) of accumulated Ci pool, and maximises the delivery of CO2 to the enzyme’s active sites via a CA localised in the lumen of trans-pyrenoidal thylakoids [1].

In the model green alga Chlamydomonas, the induction of the CCM is accompanied by in-depth transcriptional, translational, and post-translational changes, as well as modification in the subcellular localisation of several soluble proteins (Rubisco, the lumenal CA) [2]. The components of the signalling and regulatory cascade, however, are unknown. With the exception of a so-called ‘master regulator’ of the CCM and a bona fide transcription factor controlling two known CCM genes, little is known about the regulation of the CCM.

The aim of the project is to advance our fundamental understanding of CCM gene regulation in Chlamydomonas by a series of experiments combining molecular, physiological, and cytological tools. The project has two main components. Firstly, provide a detailed characterisation of existing knock-out and knock-down regulation mutant candidates. Secondly, develop tools for the analysis of cis/trans-acting elements and DNA-binding proteins involved in CO2-responsive transcriptional activation of CCM genes.

(i)         Validation and evaluation of 12 insertional mutants generated by collaborators in Stanford. These mutants are predicted to code for putative transcription factors, post-translational modification enzymes, or chaperones. Each insertion will be validated by PCR first. Then, independent transformants will be screened for impairment in Ci assimilation, via spot test and affinity for CO2 (O2 electrode).  The effect on expression of known CCM genes will be probed by qRTPCR. Finally, ultrastructural changes on chloroplast and pyrenoid in particular will be examined by transmission electron microscopy, coupled to immunugold labelling.

(ii)        To identify cis-elements involved in the transcriptional regulation of mobile CCM genes, a series of 5’-nested deletions of the region upstream of LCIB, a peri-pyrenoidal protein which acts as a putative CO2-leakage barrier, will be fused to a promoterless reporter gene. Induction/repression of the reporter gene will be quantified via qRTPCR, as well as biochemical assay or confocal microscopy (depending on the choice of the reporter gene) [3].

References: 

[1] Origins and diversity of eukaryotic CO2-concentrating mechanisms: lessons for the future. Meyer M, Griffiths H. J Exp Bot 2013, 64(3):769-86; doi: 10.1093/jxb/ers390.

[2] Dynamics of carbon concentrating mechanism induction and protein re-localisation during the dark to light transition in synchronised Chlamydomonas. Mitchell MC, Meyer M, Griffiths H. Plant Physiol 2014; doi : 10.1104/pp/pp.114.246918

 [3] CO2-responsive transcriptional regulation of CAH1encoding carbonic anhydrase is mediated by enhancer and silencer regions in Chlamydomonas reinhardtii. Kucho Ki, Ohyama K, Fukuzawa H. Plant Physiol 1999 121(4):1329-38.

Project availability: Michaelmas and Lent Term

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Testing the role of chromatin at Arabidopsis meiotic crossover hotspots

Supervisor: Dr Ian Henderson, Plant Sciences

Project abstract: Meiotic recombination is non-randomly distributed in plant genomes and is concentrated at hotspots located at gene promoters. Hotspots correlate both with epigenetic marks, such as the histone variant H2A.Z, and DNA sequence motifs. Additional chromatin modifications are known to be enriched at gene promoters, including histone H3K4 methylation, which contributes to regulation of gene expression. In this project the student will analyse meiotic recombination in mutants with altered H3K4 methylation to test the role of this modification in promoting crossovers. This will utilise techniques developed in the lab to measure crossovers at hotspot (pollen-typing) and megabase (fluorescence pollen flow cytometry) scale.


Project availability: Michaelmas and Lent Term

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Control fast or control smart? Balancing epidemiological knowledge against ease of control

Supervisor: Dr Nik Cunniffe, Plant Sciences

Project abstract: Invasive pathogens have considerable economic, political and humanitarian impacts. Diseases of crops and livestock can also have severe food security implications. Prominent recent and/or emerging examples can be drawn from diseases of both plants (e.g. citrus canker, citrus greening, UG99 wheat stem rust) and animals (e.g. foot and mouth, bluetongue). It is intuitively clear that any attempt to control invasive disease is most likely to succeed when any epidemic remains small, particularly in cases where control targets localised eradication. However, at the start of an epidemic little will be known about the dynamics of pathogen spread and few data are available, and so the impact of control strategies is difficult to assess and optimise. There is therefore an unfortunate but unavoidable trade-off between acting quickly but with limited knowledge, and acting slowly, which means more knowledge is available, but that there is also a larger problem to tackle.  Given the impacts of invasive pathogens, and the devastating expense of the types of control strategies that are typically adopted, this is an important issue.  However there are few published mathematical models targeting this question.  Our initial exploratory work has used a compartmental SIR type model to identify how optimal decision making depends on the underlying epidemiological dynamics and the availability of data characterising pathogen spread.  Extending this work will be the focus of the project, in particular to include stochastic and spatial aspects of the problem, and a more detailed consideration of the set of possible controls that are available.

Learning outcomes and skills acquired:The student will gain expertise in mathematical modelling, computer simulation and challenging models with data.  They will also gain experience of how models can affect policy.

Project availability: Michaelmas and Lent Term

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Epidemiological modelling: effects of the spatial resolution of host data on crop disease epidemics

Supervisor: Professor Chris Gilligan, Plant Sciences

Project abstract: Recurrent and emerging crop pathogens are of increasing concern worldwide, and have obvious effects on food security. Extensive modelling work using spatially-explicit, stochastic, compartmental models has been used to understand how these pathogens spread, and in turn how they might best be controlled. However, to work within reasonable timescales at the spatial scales that can be required (e.g. entire countries or even continents) these models often use the structured metapopulation paradigm. This involves sampling available spatial data on the location of host crops at a particular spatial resolution (e.g. 1 hectare squares), and treating all hosts within each square as a homogeneous whole. While this has clear benefits in terms of making large scale simulations run acceptably fast, it also undoubtedly has an effect on the spread dynamics that emerge. However, these effects have not been systematically considered. This project aims to address this, using an extremely detailed dataset concerning the spatial location of crops over the whole United Kingdom. In particular the effects of sampling host data at different spatial resolutions will be assessed, both in terms of the dynamics of the epidemic and also the optimal control strategy that would be recommended. The understanding we will gain will be an important component of our understanding of a class of mathematical models that are not only important for global food security, but are already being used to inform policy (for a representative example from the U.K. in the context of the tree disease sudden oak death see: http://www.fera.defra.gov.uk/plants/plantHealth/pestsDiseases/phytophthora/documents/openMeeting/cambridgeModelling.pdf).

Learning outcomes and skills acquired:The student will gain skills in experimental design and hypothesis testing in ecological modelling. The student will have the opportunity to learn how to use and adapt stochastic models for the spread of disease through heterogeneous populations using data for the spatial distributions of crops within the UK. S/he will learn to use the CamGrid Computer cluster, graphical techniques for summarising simulation outputs and statistical methods for model comparison. S/he will learn how models are used to inform practical control strategies for emerging epidemics,  in addition to learning fundamental techniques in modelling.

Project availability: Michaelmas and Lent Term

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Fatal attraction?  Epidemiological consequences of vector preference for plant disease

Supervisor: Dr Nik Cunniffe

Project abstract: Virus diseases of plants are most often transmitted by vectors, typically herbivorous insects that spread the pathogen as they feed on different plants. That vectors of disease can be affected by host plant infection status has been known for a number of years. Indeed, it is known that vectors can exhibit orientation preference, in which they preferentially land on infected plants, or feeding preference, in which they preferentially settle to feed on infected plants after landing, or neither, or both.  The preference of an individual vector can also depend on whether it is currently infected. Clearly these responses will affect the dynamics of disease spread. However these observations have only just started to be integrated into mathematical models. The epidemiological and evolutionary consequences are therefore not properly understood; neither is whether the responses can be manipulated to the detriment of the pathogen. Taking the recent emergence of cassava brown streak as the biggest threat to food security in East Africa as just one example, it is clear that anything offering hope of low cost control of viral disease would be a timely contribution.

Learning outcomes and skills acquired: The student will gain expertise in abstracting a complex biological system into a mathematical model. They will have the opportunity to develop deterministic and stochastic models of the plant-virus-vector interaction, and will learn how to understand the behaviour of these models via mathematical analysis and simulation.

Project availability: Lent Term (January - March 2015)

Other relevant themes: World class underpinning bioscience

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Does over-nutrition cause dementia via activating microglia?

Supervisor: Prof. Guy C. Brown

Project abstract: Microglia are brain macrophages involved in shaping, protecting and killing brain neurons in development, aging and neurodegeneration.  We have been investigating the mechanisms by which microglia become inflamed and damage neurons.  Recently we have found that inflamed microglia can phagocytose (i.e. eat) live neurons, and thereby cause neuronal death and loss. This project will investigating the signals and receptors involved in this process, and whether phagocytosis is beneficial or detrimental in particular pathologies.  The project will use cell culture, fluorescence microscopy and molecular cell biology.       Over-nutrition and diabetes have recently been found to increase the risk of dementia, but the causes are unclear.  We have found that glucose and deoxyglucose can affect microglial inflammation. This project will investigate how nutrients affect microglial activation and their ability to protect of kill neurons in culture.    Brown GC & Neher JJ (2014) Microglial phagocytosis of live neurons. Nat. Rev. Neurosci. 15, 209-216.   Neher JJ, Emmrich JV, Fricker M, Mander PK, Thery C, Brown GC (2013) Phagocytosis executes delayed neuronal death after focal brain ischemia. Proc Natl Acad Sci 110:E4098-107.     Keywords: Neuroscience, Neuroinflammation, Diabetes, Cell Death, Nutrition

Learning outcomes and skills acquired: The student will acquire skills in cell culture, biochemistry, fluorescence microscopy and molecular cell biology.

Project availability: Lent Term (January - March 2015)

Other relevant themes: Basic bioscience underpinning health, World class underpinning bioscience

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Regulation of gene expression by microRNAs as a strategy to optimise costs and benefits of disease resistance in plants.     posttranscriptional regulation, translation, effector triggered immunity, ATP binding sites, controlling endogenous RNA

Supervisor: Prof Sir David Baulcombe

Project abstract: Plants have to trade off the costs and benefits of disease resistance. To reduce the costs of disease resisance in the absence of pathogens many disease resistance mechanisms are inducible.  However inducible disease resistance will only be fully effective after an infection is established and is less effective than a mechanism that is constitutively active. Extreme resistance is imcompatible with the highest levels of fitness and in wild plants there will be a balance   We have recently described a microRNA superfamily that is a global downregulator of defense and we proposed that these RNAs mediate the trade-off in disease resistance. This project will test that hypothesis using a transient gene expression system in Nicotiana and tomato. We will design an RNA that will bind to members of the miRNA superfamily so that they are diverted away from their normal targets. We will then assemble constructs to express these “decoy” or “sponge” RNAs and assay their effects on the profile of gene expression using next generation sequencing and on disease resistance by inoculation with various pathogen.  We predict that more effective sponges will block the microRNA regulators of disease resistance and result in upregulation of disease resistance. The next stage– to test fitness costs - will unfortunately be beyond the scope of this short project.

Learning outcomes and skills acquired: Familiarity with the literature on miRNAs and disease resistance in plants.  Bioinformatics both in construct design and in the analysis of next generation sequencing data.  Plant pathology and inoculation with pathogenic fungi, oomycetes, bacteria and viruses.  Presentation of data and literature in group lab meetings.   Appreciation of the evolutionary dynamics of disease resistance

Project availability: Lent Term (January - March 2015)

Other relevant themes: IBBE, World class underpinning bioscience

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Hormone involvement in plant shape

Supervisor: Dr Siobhan Braybrook

Project abstract: Classical experiments have given us broad descriptive strokes describing major hormone action in plant development: auxins trigger root formation and cytokinins trigger shoots.  But there are more hormones to be considered, and each has very specific effects on cell growth: auxin, gibberellin, brassinosteroids, and cytokinin lead to cell growth; ABA and ethylene repress growth.  Again, these are relatively broad strokes as these hormones have different effects under different environmental conditions, for example gibberellins trigger elongation in the light but not the dark.     The aim of this project is to characterize the effect of several plant hormones on cell shape, organ growth, and cell wall mechanical properties length in Arabidopsis hypocotyls. Analyzing the effects of these hormones on growth and the cell wall in germinating seeds/hypocotyls could have profound impacts on early seedling establishment in agriculture. This project will also expose the researcher to an exciting interdisciplinary lab with engineers, biologists, physicists working together to solve biological problems.    We will manipulate hormone responses by 1) exogenous application of active compounds or inhibitors, 2) analysis of hormone reporters during growth, and 3) hormone mutant growth analysis.   Growth will be analyzed with ifra-red growth tracking, cell shape assessed by scanning electron microscopy and confocal microscopy, hormone response reporters will be imaged with confocal microscopy, and cell wall mechanics tested using an atomic force microscope. There is potential for generation of new hormone reporter constructs as well.

Learning outcomes and skills acquired: Confocal, scanning electron, and atomic force microscopy. Experience in growth tracking and analysis. Experience in hormone response reporter imaging (ABA, Auxin, GA). Learning how to assess the mechanical properties of growing biological tissues. Presentation skills developed in lab meetings and poster sessions when applicable. Experience working in an interdisciplinary lab.

Project availability: Lent Term (January - March 2015)

Other relevant themes: IBBE, World class underpinning bioscience

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Molecular Genetic Reconstructions from DNA Sequences of Past Effective Population Sizes of Animal Populations

Supervisor: Professor William Amos

Second supervisor: Professor Rhys Green

Project abstract: In recent years, the effective sizes of many animal populations have been reconstructed over periods extending hundreds of thousands of years or more back into the past using DNA sequence data. If reliable, these reconstructions are of enormous value in population ecology, population genetics and conservation science and in devising sustainable agricultural systems.  Knowledge of past population sizes in pre-agricultural times is important in interpreting alternative approaches to reconciling biodiversity conservation with food production, and therefore for achieving food security goals at least cost to wild species.  This project will examine the sensitivity of population reconstructions  to the methods used, choices of parameter values, and the validity of the underlying assumptions of the models used to obtain them. It will also analyse reconstructed effective population trajectories across sets of many species to search for possible signals left by past changes in climate and habitat extent for which there are independent data, such as those produced by glaciations. You will learn about the methods used to reconstruct past population size from DNA sequence data and test the effects upon the results of changing assumptions and using different methods and input parameter values. You will compile published population reconstructions and their accompanying data and metadata. You will compile mitochondrial DNA sequence data from public databases.  You will perform your own population reconstructions from mtDNA sequence data. You will analyse multi-species sets of population trajectories and search the results for associations with independent paleoclimate and paleovegetation time series.

Learning outcomes and skills acquired: The student will gain experience in the estimation of effective population size from DNA sequence data, population genetics models and the statistical analysis of multiple datasets. More generally they will be given a grounding in project design.

Project availability: Lent Term (January - March 2015)

Other relevant themes: World class underpinning bioscience, Food Security

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Impacts of Agricultural Production on Population Size of Wild Species using Molecular Genetic Reconstructions

Supervisor: Professor Bill Amos

Second supervisor: Professor Rhys Green

Project abstract: Knowledge of past population sizes in pre-agricultural times is important in interpreting alternative approaches to reconciling biodiversity conservation with food production in the modern world, and therefore for achieving food security goals at least cost to wild species.  In recent years, the effective sizes of many animal populations have been reconstructed over periods extending hundreds of thousands of years or more back into the past using DNA sequence data. If reliable, these reconstructions are of enormous value in population ecology, population genetics and conservation science and in devising sustainable agricultural systems.  This project will examine the sensitivity of population reconstructions to the methods used, choices of parameter values, and the validity of the underlying assumptions of the models used to obtain them. It will also analyse reconstructed effective population trajectories across sets of many species to search for possible signals left by past changes in climate and habitat for which there are independent data, such as those produced by glaciations and the spread of agriculture. You will learn about the methods used to reconstruct past population size from DNA sequence data and test the effects upon the results of changing assumptions and using different methods and input parameter values. You will compile published population reconstructions and their accompanying data and metadata. You will compile mitochondrial DNA sequence data from public databases.  You will perform your own population reconstructions from mtDNA sequence data. You will analyse multi-species sets of population trajectories and search the results for associations with independent paleoclimate and paleovegetation time series.

Learning outcomes and skills acquired: The student will gain experience in the estimation of effective population size from DNA sequence data, population genetics models and the statistical analysis of multiple datasets. More generally they will be given a grounding in project design.

Project availability: Lent Term (January - March 2015)

Other relevant themes: World class underpinning bioscience, Food Security

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