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    • 4. 发明公开
    • METHODS OF SCREENING NUCLEIC ACIDS FOR SINGLE NUCLEOTIDE VARIATIONS
    • 筛选单核苷酸变异的核酸的方法
    • EP2152915A2
    • 2010-02-17
    • EP08754474.8
    • 2008-05-14
    • Insight Genetics, Inc.
    • DAHLHAUSER, Eric, B.
    • C12Q1/68
    • C12Q1/6827C12Q1/6834C12Q2535/125C12Q2525/125C12Q2521/319C12Q2521/101
    • Disclosed are methods and compositions for detecting variation in nucleic acids. The disclosed method compares the sequence of a nucleic acid of interest with the sequence of a reference nucleic acid to sensitively identify variations between the sequence of a nucleic acid of interest and the sequence of a reference nucleic acid. The disclosed method generally involves excision and replacement of selected nucleotides in nucleic acid strands hybridized to other strands. In the method, if the excised nucleotide was mismatched with the nucleotide in the other, hybridized strand, then the replacement nucleotide will not be mismatched. If the excised nucleotide was not mismatched with the nucleotide in the other, hybridized strand, then the excised nucleotide is not replaced. This difference allows detection of variation in the nucleic acid of interest. In some forms of the method, by replacing excised nucleotides with nuclease-resistant nucleotides, strands in which excised nucleotides are replaced will be resistant to nuclease digestion while strands in which excised nucleotides are not replaced will be sensitive to nuclease digestion. By exposing the hybridizing nucleic acids to nuclease following replacement of excised nucleotides, the strands in which excised nucleotides are not replaced can be destroyed by the nuclease while strands in which excised nucleotides are replaced can be preserved. The remaining strands can then be detected and whether the strand survived nuclease digestion can be noted. Strands that survive nuclease digestion are indicative of the presence of variation in the nucleic acid of interest.
    • 公开了用于检测核酸变异的方法和组合物。 所公开的方法将感兴趣的核酸的序列与参考核酸的序列进行比较以灵敏地鉴定感兴趣的核酸的序列与参考核酸的序列之间的变化。 所公开的方法通常包括切除和置换与其他链杂交的核酸链中的选定核苷酸。 在该方法中,如果切下的核苷酸与另一杂交链中的核苷酸错配,则替换的核苷酸不会错配。 如果切除的核苷酸不与另一条杂交链中的核苷酸错配,则切除的核苷酸不被替换。 这种差异允许检测感兴趣的核酸的变异。 在该方法的一些形式中,通过用核酸酶抗性核苷酸置换被切除的核苷酸,其中被切除的核苷酸被置换的链将对核酸酶消化有抗性,而其中被切除的核苷酸不被置换的链将对核酸酶消化敏感。 通过在交换的核苷酸置换后将杂交核酸暴露于核酸酶,其中被切除的核苷酸不被置换的链可被核酸酶破坏,而被置换的核苷酸置换的链可被保存。 然后可以检测到剩余的链,并且可以注意到链是否存在核酸酶消化。 存活核酸酶消化的链表明目标核酸存在变异。