HIGHLY SENSITIVE METHOD FOR DETECTING LOW FREQUENCY MUTATIONS
    28.
    发明公开
    HIGHLY SENSITIVE METHOD FOR DETECTING LOW FREQUENCY MUTATIONS 审中-公开
    高SENSIBLES检测方法的突变低

    公开(公告)号:EP2814985A2

    公开(公告)日:2014-12-24

    申请号:EP13749733.5

    申请日:2013-02-13

    IPC分类号: C12Q1/68

    摘要: The disclosed edge-blocker oligonucleotide based AS-NEPB-PCR method amplifies allele specific DNA (or RNA) while dramatically blocking amplification of wild type (WT) DNA (or RNA). The AS-NEPB-PCR design allows ready modification of an existing PCR reaction setup with an edge-blocker oligonucleotide together with an allele specific primer complementary to the mutant sequence to achieve allele specific amplification. The method simplifies assay optimization procedures and achieved sensitivity sufficient to detect a signal present at 0.1% level with close to 100% specificity, which is useful in detecting SNP or genetic mutations. The method was used to detect three different genetic mutations in cancer, in KRAS, BRAF, and EGFR, with three different types of modified edge-blocker oligonucleotides (phosphate, inverted dT and amino-C7). It was possible to detect one copy of mutant DNA in 1000-copy of normal DNA background of a heterogeneous sample, and was far more sensitive than the other blocking method.