The primary objective of this proposal is to identify and clone genes involved in the processes of calcium carbonate biomineralization and coccolithogenesis in the marine coccolithophorid, Emiliania huxleyi. Biomineralized skeletons have been targeted as having important applications in biomedical materials chemistry for the construction of artificial bone in humans, scaffolding supports in tissue engineering, complement activation enhancement, artificial dental root construction, and biomedical implant technology. They also have potential significance as a material source for light weight ceramics, catalyst supports, and robust membranes for high temperature separation technologies. While a general understanding of some of the ecophysiological aspects of calcification and coccolithogenesis in E. huxleyi has been obtained, little information is available on the molecular mechanisms that govern these processes. The research approach in this proposal is designed to test the following hypotheses: 1) the production and assembly of coccoliths in E. huxleyi requires the expression of specific genes and gene products; 2) specific genes and gene products are responsible for the regulation of coccolith production during various stages of growth in response to environmental stimuli; and 3) E. huxleyi will be amenable to genetic transformation. Initial work will employ chemical mutagenesis and classical genetic techniques to identify and clone genes essential to calcification and coccolithogenesis in E. huxleyi. A genetic transformation system will also be developed utilizing microparticle bombardment and employing vectors which either confer resistance to antibiotics, or that contain autotrophic genes as selectable markers. The expression of genes essential will enable alternative approaches to isolate and clone genes, such as differential display and subtractive hybridization, to be employed. The results of these studies will contribute towards understanding the design principles of biomineralized CaCO3 structures, and thus will provide models for novel synthesis and processing strategies for producing materials for biomedical applications.
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