SBE-TAGS: an array-based method for efficient single-nucleotide polymorphism genotyping.

ABSTRACT

Generating human single-nucleotide polymorphisms (SNPs) is no longer a rate-limiting step for genetic studies of disease. The number of SNPs in public databases already exceeds 200,000, and the total is expected to exceed 1,000,000 within a year. Rather, progress is limited by the inability to genotype large numbers of SNPs. Current genotyping methods are suitable for studying individual loci or at most a handful at a time. Here, we describe a method for parallel genotyping of SNPs, called single base extension-tag array on glass slides, SBE-TAGS. The principle is as follows. SNPs are genotyped by single base extension (SBE), using bifunctional primers carrying a unique sequence tag in addition to a locus-specific sequence. Because each locus has a distinct tag, the genotyping reactions can be performed in a highly multiplexed fashion, and the resulting product can then be "demultiplexed" by hybridization to the reverse complements of the sequence tags arrayed on a glass slide. SBE-TAGS is simple and inexpensive because of the high degree of multiplexing and the use of an easily generated, generic tag array. The method is also highly accurate we genotyped over 100 SNPs, obtaining over 5, 000 genotypes, with approximately 99% accuracy.