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    • 10. 发明授权
    • Method of extracting particular nucleic acid fragment
    • 提取特定核酸片段的方法
    • US5252724A
    • 1993-10-12
    • US700694
    • 1991-05-16
    • Toshihiko KishimotoShin-Ichiro NiwaAtsushi Uno
    • Toshihiko KishimotoShin-Ichiro NiwaAtsushi Uno
    • C07H1/00C12N15/10C12Q1/68
    • C07H1/00C12N15/10
    • A method of extracting a particular nucleic acid fragment containing a nucleic acid sequence of interest from a nucleic acid or nucleic acids mixture, comprising the steps of:(1) digesting the nucleic acid or nucleic acids mixture with restriction enzymes to obtain a mixture of nucleic acid fragments, said restriction enzymes consisting of (A) two different enzymes capable of producing the particular nucleic acid fragment bearing predetermined and distinct restriction ends on its 5' and 3' terminals and (B) one or more restriction enzymes different from (A), for which the particular nucleic acid fragment contains no relevant restriction sites;(2) preparing two distinct DNA linkers capable of binding to the respective restriction ends of the particular nucleic acid fragment;(3) allowing the linkers to react with the mixture of nucleic acid fragments;(4) subjecting the resulting reaction mixture to the first hybridization with an immobilized probe complementary to one of the linkers;(5) isolating the hybridized nucleic acid fragment from the probe;(6) subjecting the isolated nucleic acid fragment to the second hybridization with an immobilized probe complementary to the other linker; and(7) isolating the hybridized nucleic acid fragment from the probe.
    • 一种从核酸或核酸混合物提取含有目标核酸序列的特定核酸片段的方法,包括以下步骤:(1)用限制酶消化核酸或核酸混合物以获得核酸或核酸混合物的混合物 所述限制酶由(A)能够产生其5'和3'末端具有预定和不同限制性末端的特定核酸片段的两种不同酶,(B)不同于(A)的一种或多种限制酶, 特定核酸片段不含有相关限制性位点; (2)制备能够结合特定核酸片段的相应限制性末端的两个不同的DNA接头; (3)允许接头与核酸片段的混合物反应; (4)使所得到的反应混合物与固定化的探针与其中一个接头互补的第一次杂交; (5)从探针中分离杂交的核酸片段; (6)使分离的核酸片段与另一个接头互补的固定化探针进行第二次杂交; 和(7)从探针中分离杂交的核酸片段。