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Cellular analysis of gene expression during development pathogen interactions and abiotic stress in plants

Reference	BB/C51103X/1
Principal Investigator / Supervisor	Dr David Leader
Co-Investigators / Co-Supervisors	Dr Jacqueline Barker, Dr Hector Cabrera Poch, Dr Rosane Curtis, Dr Ruth Gordon-Weeks, Dr Konstantin Kanyuka, Dr Andrew Phillips, Professor John Pickett, Professor Peter Shewry
Institution	Rothamsted Research
Department	Plant Biology & Crop Science
Funding type	Research
Value (£)	56,738
Status	Completed
Type	Research Grant
Start date	05/01/2005
End date	04/10/2005
Duration	9 months

Abstract

Laser capture microdissection (LCMD) is an efficient and versatile technique for analysing gene expression at the cellular level. We propose to utilise this technique using a PALM microbeam IP laser dissection microscope to investigate the spatial regulation of gene expression in several biological systems being studied within different groups at Rothamsted Research. (1) Wheat grain development. LCMD will be used to analyse differences in gene expression within the endosperm of wheat different storage protein genes during the formation of the starchy endosperm. It will also be used to undertake expression microarray studies to identify genes which are specifically associated with cell differentiation during wheat grain development. (2) Virus transmission by Polymyxa graminis in wheat and barley. LCMD will be used to study biology, taxonomy and evolution of agriculturally important but poorly understood zoosporic microorganisms, Polymyxa graminis, which thrives in roots of cereal plant species and has a unique ability to serve as a vector for a number of plant viruses, causal agents of mosaic diseases. This equipment will also aid identification of the mechanism of resistance to the P. graminis-transmitted soil-borne cereal mosaic virus in winter wheat, and will help to answer several other important fundamental questions on the P. graminis (vector) ¿ virus relationship. (3) Mechanisms of heavy metal tolerance and hyperaccumulation in plants. LCMD will be used to sample groups of different cell types throughout the metal transport pathway of -/+ metal-treated P. vittata (arsenic) and T. caerulescens (cadmium and zinc). Cell-type specific samples will be used for RNA extraction and subsequent gene expression studies by real-time RT-PCR, as well as for probes for cDNA library screening. Comparisons of cell-type specific gene expression between high- and low-Cd accumulating ecotypes and transgenic lines of T. caerulescens will also be examined. (4) Analysis of genes regulating host pathogen interactions between potatoes and potato cyst nematodes. LCMD will enable the isolation of nematode parasitism genes from the specific plant nematode cells involved in triggering and maintenance of the host feeding site. Similarly, host genes that are differentially regulated in these feeding sites can be isolated and further characterised by precisely excising these cellular structures out of the rest of the roots by LCMD. (5) Tissue specific transcriptome analysis of cereal responses to aphid feeding and allelopathic agents. LCMD will be used to analyse tissue specific changes in gene expression that lead to the local and systemic accumulation of defence proteins and metabolites in cereals in response to aphids and allelopathic compounds. The epidermis and vascular tissue will be excised by LCMD and gene expression will be analysed by PCR and cDNA-ALFP. (6) Hormones and the control of plant growth and development. LCMD will be used to improve the spatial resolution of gibberellin biosynthesis and signalling in two experimental systems. In root growth, LCMD will be used to map expression of the GA biosynthetic and response genes to determine the extent of movement of intermediates and bioactive GAs; the regulation of leaf shape in tomato will be studied by mapping the interaction of KNOX meristem identity genes with cytokinin and GA biosynthetic genes.

Summary

unavailable

Committee	Closed Committee - Plant & Microbial Sciences (PMS)
Research Topics	X – not assigned to a current Research Topic
Research Priority	X – Research Priority information not available
Research Initiative	Research Equipment Initiative 2004 (RE4) [2004]
Funding Scheme	X – not Funded via a specific Funding Scheme

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