Comparison of PrASE and Pyrosequencing for SNP Genotyping

The assay is automated for 48 genotyping reactions in parallel followed by a tag-microarray detection system. A script automatically visualizes the results in cluster diagrams and assigns the genotypes. Ten polymorphic positions suggested as prothrombotic genetic variations were analyzed with Pyrosequencing and PrASE technologies in 442 samples and 99.8 % concordance was achieved. In addition to accuracy, the robustness and reproducibility of the technique has been investigated.The results of this study strongly indicate that the PrASE technology can offer significant improvements in terms of accuracy and robustness and thereof increased number of typeable SNPs.It is now a common belief that single nucleotide variations in the human genome are responsible for influencing traits such as differences in drug metabolism and disease risk. These variations are referred to as single nucleotide polymorphisms (SNPs) and several large-scale technologies have recently been developed for scoring of thousands of SNPs and approaching whole-genome genotyping [1-5].However, for smaller scale projects where potential genes are already known, technologies for genotyping of many samples instead of SNPs and in addition retain high accuracy and throughput, are more attractive compared to assays that are cost effective per SNP. A flexible choice of SNPs is also important instead of a pre-defined set of SNPs. There are several technologies already used in academic contexts but the earliest paralleled assays relied upon hybridization of short allele-specific probes to the target DNA [6,7]. However, improvements in microarray-based technologies in terms of accuracy have been achieved by enzymatic means [8-10]. One of these technologies involves allele-specific extension (ASE) which utilizes the ability of DNA polymerase to distinguish matched and mismatched 3'-termini of primers. However, a number of reports have shown that some mismatched 3'-ends can be elongated, giving false positive sig