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JMD 2003, Vol. 5, No. 4
Copyright © 2003 American Society for Investigative Pathology & Association for Molecular Pathology

Comparison of GenFlex Tag Array and Pyrosequencing in SNP Genotyping

Daniel C. Chen*, Janna Saarela*{dagger}, Ilpo Nuotio*, Anne Jokiaho*, Leena Peltonen*{dagger}{ddagger} and Aarno Palotie*§

From the Department of Human Genetics, * David Geffen School of Medicine at UCLA, Los Angeles, California; the Department of Molecular Medicine, {dagger} National Public Health Institute, Helsinki, Finland; the Department of Medical Genetics, {ddagger} University of Helsinki, Finland; the Department of Pathology and Laboratory Medicine, § David Geffen School of Medicine at UCLA, Los Angeles, California; and the Department of Clinical Chemistry and the Finnish Genome Center, University of Helsinki, Finland

With the completion of the Human Genome Project, over 2 million sequence-verified single nucleotide polymorphisms (SNPs) have been deposited in public databases. The challenge has shifted from SNP identification to high-throughput SNP genotyping. Although this has had little impact on molecular diagnostics, it provides the potential for future molecular diagnostics of complex traits to include SNP profiling. Accordingly, efficient, accurate, and flexible SNP genotyping are needed. In addition, the drive for low cost has pushed genotyping reactions toward multiplexing capability. We compared two SNP genotyping techniques: Affymetrix GenFlex Tag array and Pyrosequencing. The reference method was a well-established, solid-phase, single nucleotide extension reaction technique based on tritium detection. Fourteen SNPs were selected from the fine mapping project of a multiple sclerosis locus on chromosome 17q. Using all three techniques and the reference method, the SNPs were analyzed in 96 related individuals. Without extensive optimization, we successfully genotyped 11 of 14 SNPs with both GenFlex and Pyrosequencing. Our study suggests that the Pyrosequencing technique provides higher accuracy between the two systems which is most likely due to the single-stranded template in the extension reaction. Thus, Pyrosequencing has potential for diagnostic applications. Pyrosequencing, however, is not optimal for large SNP profiling analyses wherein multiplexing potential is an advantage.




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