The ability to determine the genotype of an individual tumor will have tremendous impact on both determining a patient's prognosis and on the design of therapeutic regimes. The purpose of the proposed project is to develop an easier and more cost effective approach to amplification of nucleic acid sequences and the detection of genetic anomalies in human tissue. The goal is to perform the amplification reaction directly on tissue prints without prior isolation of the nucleic acids. Fresh tumor tissue collected directly following surgery or biopsy will be used for generating tissue prints. Preliminary data demonstrates that several matrix materials can be used to immobilize biological specimens for successful direct amplification of nucleic acids. These and other matrix materials will be tested for the efficiency of cell transfer from a solid tissue print. The conditions for amplification of single-copy human genes, mRNA and the detection of point mutations will be optimized using tissue prints directly in the enzymatic amplification reactions. Detection of amplified DNA and mRNA products and the discrimination of wild-type and mutant genes will be performed using DNA microprobe arrays. Long range goals include integration of sample collection and processing into a single device and an automated platform.
Automated diagnostic tests for gene mutations and aberrant gene expression in tumor tissue will improve patient care as well as reduce the cost and speed at which genetic testing can be done.
