Quantitative real-time PCR has become the standard for clinical quantitation of infectious agents such as HIV, herpesviruses, West-Nile virus and others. It is used extensively in basic research everywhere. It is currently the method of choice to confirm microarray data or data obtained from quantitative deep sequencing experiments. We developed real-time QPCR arrays for human viruses, for targeted profiling of cell signaling pathways such as NFkappaB target genes and for human microRNAs. The PI's group has become the real-time QPCR core facility for the UNC microbiology community and the NCI-designated Lineberger comprehensive cancer center (LCCC). The dedicated real-time QPCR core is located in the Department of Microbiology and unlike other units operates under BSL-2 containment to be able to work with infectious agents and infectious samples, including those that require worker immunization and/or federal clearance. The core consists of a Agilent Bioanalyzer for QC of input RNA and post QPCR analysis, and Tecan general pipetting robot and integrated LC480 QPCR instrument. This allows us to load 384-well plates with 3 ?mu?l reaction volume and to run the QPCR instrument fully automated for 24 hours a day. Our main reason for choosing a plate-based system compared to e.g. nanofluidics or microarray systems, was that even a 384-well plate allow post-PCR processing, which present nanofluidics devices do not. In addition to the quantitative data of each experiment, the physical product is available for downstream applications such as QC, sequencing or cloning. In sum, we have a published track record in the development, use and data analysis of targeted real-time QPCR arrays;we will serve as a core for the PPG;we the necessary infrastructure in place already, such that only operating costs and personal need to be supported.
Molecular biology and cancer research move towards high throughput experimental designs. The core will enable academic research to conduct quantitative measurements of gene expression and human viruses with unprecedented accuracy and a throughput that was previously only available in industrial settings.
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