MUTATION AND SNP IDENTIFICATION WITHOUT SEQUENCING

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The aim of this project is to develop a highly refined method of allele/mutation identification based on the use of Immobilized Mismatch Binding Protein (IMBP) and short (20-30mer) synthetic oligonucleotides. The use of short oligos with IMBP allows the simultaneous, but independent examination of several mutant or polymorphic sites in a PCR amplicon and with a single PCR amplification. The method avoids problems of PCR errors and allows long amplicons, which heretofore have created background problems with IMBP. Kits will be prepared to identify alleles at three codons of the sheep prion protein gene related to scrapie susceptibility and to precisely identify a collection of key mutations in the p53 tumor suppressor gene. The technology has immediate applications in clinical diagnostics and genomics and can largely eliminate the need to use sequencing for identification of known alleles/mutations or polymorphisms. PROPOSED COMMERCIAL APPLICATIONS: Commercial applications of a short oligo method of IMBP mutation/polymorphism detection include clinical diagnostics, where the method can largely replace sequencing for identification of known mutations. In addition, the method will be a major contributor to genetic studies involving polymorphism identification and distribution as well as studies of human sequence variation and the discovery of disease- associated genes.