Sensitive Sequencing Method for KRAS Mutation Detection by Pyrosequencing

摘要: Both benign and malignant tumors represent heterogenous tissue containing tumor cells non-neoplastic mesenchymal inflammatory cells. To detect a minority of mutant KRAS alleles among abundant wild-type alleles, we developed sensitive DNA sequencing assay using Pyrosequencing, ie, nucleotide extension with an allele quantification capability. We designed our Pyrosequencing for use whole-genome-amplified from paraffin-embedded tissue. Assessing various mixtures cell lines line, found that mutation detection rates were superior to dideoxy sequencing. In addition, proved in the mutations colon cancer normal as well pancreatic cancers. Quantification by was precise useful validation, monitoring, quality assurance. Our method is simple, robust, sensitive, limit approximately 5% alleles. It particularly applicability this amplified whole-genome amplification technique provides expanded source large-scale studies.