PE Biosystems TaqMan Quantitative PCR Model 7700

Scripps Research Institute, La Jolla CA

Investigators

Abstract

DESCRIPTION: (Investigator?s Abstract): The ability to measure gene expression levels has until recently required laborious and time consuming, or poor quantitative methods. The recent development of the "TaqMan" PCR-based system (PE Biosystems Real-time Quantitative PCR, Model 7700 Sequence Detector) allows for rapid analysis and accurate quantitation of expressed genes, as well as high throughput and multiple replicates. For example, mRNA levels for a gene of interest can be determined by northern blotting. This method is very time-consuming, due to running and blotting gels, hybridization, and washing steps and in time taken for the exposure to film or phosphorimager plate. It is difficult to analyze multiple samples and if the experimental and control genes are similar in size, a comparison to a housekeeping gene cannot always be done at the same time as the experimental exposure. The advent of the polymerase chain reaction (PCR) now allows for much more rapid analysis of gene expression, and for analysis of multiple samples. However, accurate quantitation is very difficult because of the different phases of the PCR reaction. The "Taqman" system measures expression using PCR with specific fluorescent dyes attached to a probe for the gene of interest, such that accurate quantitation of PCR product is made using fluorescence resonance energy transfer between the fluors. All tubes in a 96-sample plate can be analyzed in the same experiment, with real-time quantitative data for each cycle of PCR. This allows very accurate quantitation relative to a reference sample. This instrument represents the state of the art in real-time PCR quantitation, and is the optimal instrument for a multi-user environment such as this department, with 59 full-time staff members plus over 100 postdoctoral fellows. This instrument will be very important to the success of the projects of at least the eighteen research groups that are represented in this application.