Improved methodologies for the preservation, isolation, and characterization of mRNA are needed. Existing RNA isolation technologies are sufficient for low-resolution analysis such as northern hybridization. However, construction of quality cDNA libraries and the analysis of differential gene expression using high-resolution techniques are both sensitive to small variations in the integrity of sample RNA. Frequently, these variations are caused by sub-optimal handling and can be minimized by optimizing RNA isolation. However, variations in RNA quality are sometimes intrinsic to the samples being analyzed. For example, necrotic cells within tumor biopsies contain degraded RNA. In either case, it is essential that such variations be identified prior to the expense of high-resolution analysis using labor intensive techniques. We propose to develop a quantitative assay for RNA integrity to be used to pre-qualify RNA samples prior to use in cDNA library construction or analysis by high resolution techniques. We also wish to develop guidelines to relate intactness of RNA to suitability for use in these techniques. Such information will be an invaluable resource for both academic and industrial laboratories. Finally, we propose to use this assay for a systematic optimization of RNA isolation technologies, with the aim of maximizing the intactness of recovered RNA.
Ambion, a leading supplier of products for RNA quantitation and analysis, intends to market a quantitative assay for RNA integrity, 'Certified Quality' RNA, and improved RNA isolation systems.
