Grant: Research Tools and Instruments - Category 1 (<$150,000)

The present application requests a high-resolution Chemiluminescence detection system for the quantitative analysis of expression of proteins (1D gels), DNA and RNA using chemiluminescence and fluorescence, and for colony counting. The equipment is requested by 2 senior investigators (J. Lafond, D. Averill-Bates) and 1 junior investigator (B. Barbeau), whose NSERC research programs all entail molecular and metabolic investigations requiring quantitative analyses of expression of proteins and nucleic acids in mammalian cells and tissues. The NSERC research programs aim to understand: (1) the signalling mechanisms underlying the effects of a cholesterol-rich diet on metabolism of maternal lipids in a model of pregnant hypercholesterolemic rabbit; (2) the implication of envelope proteins derived from human endogenous retroviral sequence (HERV) in development of placenta; and (3) to identify the cellular and molecular mechanisms that regulate cellular stress responses (e.g. oxidative stress) in mammalian cells. The requested equipment is recent high-resolution (4.2 megapixels) technology that is equipped with a high sensitivity CCD camera, and software for acquisition, analysis and image enhancements, and for fluorescence and biofluorescence imaging. This equipment is essential in each of these projects for high-sensitivity detection and quantitative analysis of expression, intracellular translocation and phosphorylation of a multitude of proteins involved in cell signalling pathways, by Western blotting. Other applications include quantification of protein expression by Western blots following knock-down using siRNA technology or overexpression by recomplementation. The Molecular imager will be ideal for quantification of changes in mRNA and protein expression in response to cellular stress, and oxidative stress-induced protein damage by electrophoresis with fluorescently-labelled probes. The acquisition of a high-resolution Molecular imager is essential to ensure high quality and competitive research at the Canadian and international levels, and training of at least 100 graduate students by NSERC-funded investigators in Biochemistry, Cell and Molecular Biology.