To accomplish the stated goals of the Human Genome Project and to reap the fruits of this major undertaking, the complete sequence of a composite human genome will have to be deciphered by the year 2005 and a genetic map composed of a dense set of single nucleotide polymorphism markers must be developed for complex genetic trait analysis. There is general agreement that the technology is in place for large-scale sequencing but that sequence- ready large-insert clone retrieval will soon become the bottleneck in any significant large-scale sequencing project. Similarly, a new NHGRI initiative will undoubtedly lead to the development of a dense set of SNP markers for the human genome but it is unclear how one can best utilize these markers in population studies that require genotyping thousands of individuals with thousands of markers. In this proposal, we plan to develop two simple, flexible, homogeneous methods for DNA analysis based on fluorescence polarization detection. Specifically, we aim to (1) develop a four-color DNA genotyping method based on fluorescence polarization detection and primer extension reactions; (2) develop a multi-color PCR detection method based on fluorescence polarization detection and the 5'-nuclease """"""""TaqMan"""""""" assay for both library screening and for allelic discrimination. Once developed, these methods will facilitate clone recovery for sequencing projects and for flexible SNP genotyping in population studies. As such, they will help define the genetic factors associated with common diseases with complex inheritance patterns and the relationship between genetic and environmental influences on human health and disease.
