There is increasing evidence that, when located in cis, specific combinations of single nucleotide polymorphisms (SNPs) affect an individual's susceptibility to disease or treatment response. Methods that are suitable for haplotyping in the clinic are needed for the benefits of these results to reach patients. Computational methods for determining haplotypes are prone to errors, whereas direct haplotyping procedures are time consuming and expensive, or cannot be applied to distant SNPs. Now, Mitra et al. present an elegant new haplotyping method that overcomes these limitations. Indeed, they show that it can be used to successfully determine the genotype and phase of SNPs that are separated by as much as 45 kb.

Genotyping of two SNPs in a single DNA sample. In the reaction for the first SNP (a), green signals indicate thymine residues and red signals indicate adenine residues; for the second SNP (b), green indicates thymine and red indicates cytosine. Haplotypes can be determined when the signal from two genotyping reactions overlaps (white circles). Image courtesy of R. Mitra, Washington University School of Medicine.

The authors immobilized a dilute sample of patient DNA on a microscope slide in a thin acrylamide gel, which contains all the reagents necessary for a polymerase chain reaction (PCR). Two primer pairs incorporated in the gel allowed two SNPs of interest to be amplified. The PCR products — referred to as polonies — are immobile because the primers are modified such that one strand of the amplified DNA is covalently linked to the acrylamide matrix. So, DNA amplified by different primer pairs, but from a single DNA molecule, co-localizes. The SNPs are genotyped, one at a time, by a single base-pair extension method that uses fluorescently-labelled oligonucleotides. After each genotyping reaction the slide is scanned and the genotype of each polony is recorded digitally. The images from different reactions are merged and a computer algorithm identifies haplotypes based on the genotypes of overlapping polonies.

This is a promising method for SNP haplotyping. It has the potential to be used in the clinic, as a buccal swab provides sufficient DNA to make one slide, which can be reused for multiple genotyping reactions. This tool will also be useful for researchers interested in characterizing genetic variation, as it can be used to determine allele and haplotype frequencies in a population from pooled DNA samples.