Research Project
6294190
APPLICANT'S DESCRIPTION: Evaluating gene expression levels is crucial for understanding cancer biology and identifying prognostic indicators of cancer. A need exists for a mRNA quantification assay that combines superior sensitivity with the simplicity and versatility that are not available from reverse transcriptase-polymerase chain reaction (RT-PCR) or current transcription-mediated amplification systems. The Split Promoter Amplification Reaction (SPAR) and similar methods provide this unique combination of attributes. Here, our goal is to evaluate the feasibility of using a novel, isothermal, linear amplification technology for quantifying specific mRNAs in complex RNA samples. Our specific aims will (1) develop an assay with optimal sensitivity and specificity for quantification of a specific mRNA in a total RNA sample and (2) develop and optimize assays for quantification of multiple mRNAs in a total RNA sample (multiplexing). We will produce a fast and inexpensive assay system, for detecting and quantifying oncogenes and other genetic markers for cancer, that is both sensitive and quantitative. Phase I will determine feasibility of a product that will have high utility for cancer researchers who are quantifying gene expression. The system will be adaptable for gene quantification in numerous fields including immunology, development, and infectious diseases and will be applicable in research and diagnostics. PROPOSED COMMERCIAL APPLICATION: Ambion, a leading supplier of products for RNA analysis and quantification, will initially market a kit for the sensitive and specific detection and quantification of cancer genes. The technology will be applicable for quantifying low levels of RNA in any research field including infectious diseases, cancer research, immunology, developmental biology, and molecular biology. The research will lead to new methods and applications of transcription-based amplification. Phase II/III will see the development of diagnostic assays.
