Coupled polymerase chain reaction-restriction-endonuclease digestion-ligase detection reaction process
    1.
    发明授权
    Coupled polymerase chain reaction-restriction-endonuclease digestion-ligase detection reaction process 有权
    耦合聚合酶链反应 - 限制性内切酶消化 - 连接酶检测反应过程

    公开(公告)号:US07014994B1

    公开(公告)日:2006-03-21

    申请号:US09528014

    申请日:2000-03-17

    IPC分类号: C12Q1/68 C12P19/34 C07H21/04

    摘要: The present invention provides a method for identifying one or more low abundance sequences differing by one or more single-base changes, insertions, or deletions, from a high abundance sequence in a plurality of target nucleotide sequences. The high abundance wild-type sequence is selectively removed using high fidelity polymerase chain reaction analog conversion, facilitated by optimal buffer conditions, to create a restriction endonuclease site in the high abundance wild-type gene, but not in the low abundance mutant gene. This allows for digestion of the high abundance DNA. Subsequently the low abundant mutant DNA is amplified and detected by the ligase detection reaction assay. The present invention also relates to a kit for carrying out this procedure.

    摘要翻译: 本发明提供了一种用于从多个靶核苷酸序列中的高丰度序列鉴定一个或多个不同于一个或多个单碱基变化,插入或缺失的低丰度序列的方法。 使用高保真聚合酶链反应模拟转化,通过最佳缓冲条件促进高丰度野生型序列,在高丰度野生型基因中产生限制性内切核酸酶位点而不是在低丰度突变基因中产生限制性内切核酸酶位点。 这样可以消化高丰度DNA。 随后,通过连接酶检测反应测定扩增和检测低丰度突变体DNA。 本发明还涉及用于实施该方法的试剂盒。