Robert J. Lipshutz

Single-nucleotide polymorphisms (SNPs) are the most frequent type of variation in the human genome, and they provide powerful tools for a variety of medical genetic studies. In a large-scale survey for SNPs, 2.3 megabases of human genomic DNA was examined by a combination of gel-based sequencing and high-density variation-detection DNA chips. A total of 3241 candidate SNPs were identified. A genetic map was constructed showing the location of 2227 of these SNPs. Prototype genotyping chips were developed that allow simultaneous genotyping of 500 SNPs. The results provide a characterization of human diversity at the nucleotide level and demonstrate the feasibility of large-scale identification of human SNPs.

As the Human Genome Project and related efforts identify and determine the DNA sequences of human genes, it is important that highly reliable and efficient mechanisms are found to access individual genetic variation. It is only through a greater understanding of genetic diversity that the true benefit of the Human Genome Project will be realized. One approach, hybridization to high-density arrays of oligonucleotides, is a fast and effective means of accessing this genetic variation. Light-directed chemical synthesis has been used to generate miniaturized, high-density arrays of oligonucleotide probes. Application-specific oligonucleotide probe array designs have been developed for the rapid screening of characterized genes. Dedicated instrumentation and software have been developed for array hybridization, fluorescence detection and data acquisition and analysis. In a specific and challenging application, oligonucleotide probe arrays have been used to screen the reverse transcriptase and protease genes of the highly polymorphic HIV-1 genome to explore genetic diversity and detect mutations conferring resistance to antiviral drugs. Results from this application strongly suggest that oligonucleotide probe arrays will be a powerful tool for rapid investigations in sequence checking, pathogen detection, expression monitoring and DNA molecular recognition.