Pyrosequencing for genotyping and SNP Analysis

Pyrosequencing technology for genotyping and SNP analysis

Most importantly, Pyrosequencing technology, which is a direct DNA sequencing technique, gives unambiguous genotyping results as well as some redundant sequence information beyond the SNP position, which serves as a valuable internal control, obtained for each sample.

(Extracted from Nilsson TK, Johansson CA. A novel method for diagnosis of adult hypolactasia by genotyping of the -13910 C/T polymorphism with Pyrosequencing technology. Scand. J. Gastroenterol. 2004; 39:287–290.)

Pyrosequencing technology presents SNPs in the context of the surrounding sequence. This guarantees that each individual analysis is correct, without the need for other controls. In a clinical context, this level of certainty is highly valuable, especially as the genetic methods developed in today’s research mature into tomorrow’s genetic tests.

pyrogram genotyping Pyrogram traces of three individuals, showing 3 different genotypes of adult hypolactasia. The five peaks on the right act as the internal control, ensuring that the assay is correct. From top to bottom, the analytical results AA, GA and GG, respectively, corresponding to the genotypes TT, CT and CC. The CC genotype is associated with genetically determined malabsorption of lactose. CT heterozygotes have about half the enzyme activity of subjects homozygous for the lactase persistence allele TT. (From Nilsson & Johansson, 2004)

Pyrosequencing technology gives an assay for any genetic marker

We were told by a major supplier of genotyping technology that an assay for trisomy couldn't be achieved in a quantitative manner. We turned to Pyrosequencing, and the result looks beautiful. Pyrosequencing is certainly a facilitator for new and useful genetic analyses in research.

Dr. Neil Lamb, Director, Center for Medical Genomics, Emory University, Atlanta, GA

An estimated 20-30% of SNPs in large-scale SNP studies are rejected because of failure of the genotyping assay. Pyrosequencing technology can provide functional assays for most, if not all, difficult SNPs and mutations.

Pyrosequencing technology has an unbeaten record of success for assaying any polymorphism or mutation. Assays can be designed with great flexibility in primer placement. Pyrosequencing assay design is both flexible and simple to do. QIAGEN provides Assay Design Software to support users.

Pyrosequencing technology therefore gives researchers full freedom to explore genetic markers unhindered by limitations in technology, based on robust assays that are easily translated to standard methods in the research community, and homebrew assays by CLIA-certified labs.