A new primer construction technique that effectively increases amplification of rare mutant templates in samples

Abstract

Background: In personalized medicine, companion diagnostic tests provide additional information to help select a treatment option likely to be optimal for a patient. Although such tests include several techniques for detecting low levels of mutant genes in wild-type backgrounds with fairly high sensitivity, most tests are not specific, and may exhibit high false positive rates. In this study, we describe a new primer structure, named 'stuntmer', to selectively suppress amplification of wild-type templates, and promote amplification of mutant templates. Results: A single stuntmer for a defined region of DNA can detect several kinds of mutations, including point mutations, deletions, and insertions. Stuntmer PCRs are also highly sensitive, being able to amplify mutant sequences that may make up as little as 0.1% of the DNA sample. Conclusion: In conclusion, our technique, stuntmer PCR, can provide a simple, low-cost, highly sensitive, highly accurate, and highly specific platform for developing companion diagnostic tests.