公开(公告)号：EP0530998B1

公开(公告)日：1996-11-13

申请号：EP92307441.3

申请日：1992-08-13

申请人： MICROGENICS CORPORATION

发明人： Eisenbeis, Scott J.

IPC分类号： C12Q1/68 ,  C12Q1/70 ,  G01N33/535

CPC分类号： C12Q1/6816 ,  C12Q2565/107 ,  C12Q2563/125

公开(公告)号：EP0530998A1

公开(公告)日：1993-03-10

申请号：EP92307441.3

申请日：1992-08-13

申请人： MICROGENICS CORPORATION

发明人： Eisenbeis, Scott J.

IPC分类号： C12Q1/68 ,  C12Q1/70 ,  G01N33/535

CPC分类号： C12Q1/6816 ,  C12Q2565/107 ,  C12Q2563/125

摘要： The invention relates to a method for detection of a specific nucleic acid sequence which comprises
   forming a reaction mixture by combining

(1) a sample suspected of containing a nucleic acid;
(2) a probe/enzyme donor polypeptide conjugate comprising

(a) an enzyme donor polypeptide sequence comprising a β-galactosidase fragment; and
(b) a single-stranded oligonucleotide sequence attached to (a) and capable of hybridizing with said nucleic acid;

(3) an enzyme acceptor polypeptide capable of forming an active enzyme upon complementation with said enzyme donor fragment; and
(4) a substrate for β-galactosidase; and

   detecting hybridization of said probe/enzyme donor conjugate to said sample nucleic acid to form a double strand-specific sequence by determining the amount or rate of enzyme activity on said substrate in said reaction mixture. The method can also include a "proof reading" function by incubating the hybridized probe with at least one double-strand-specific, sequence-specific restriction endonuclease.
Novel kits for use in carrying out the method are also included.

摘要翻译： 本发明涉及一种用于检测特异性核酸序列的方法，其包括通过组合（1）怀疑含有核酸的样品形成反应混合物; （2）探针/酶供体多肽缀合物，其包含（a）包含β-半乳糖苷酶片段的酶供体多肽序列; 和（b）与（a）连接并能够与所述核酸杂交的单链寡核苷酸序列; （3）能够在与所述酶供体片段互补时能够形成活性酶的酶受体多肽; 和（4）β-半乳糖苷酶的底物; 以及通过确定所述反应混合物中所述底物上的酶活性的量或速率来检测所述探针/酶供体缀合物与所述样品核酸的杂交以形成双链特异性序列。 该方法还可以通过将杂交探针与至少一个双链特异性的序列特异性限制性内切核酸酶孵育来包括“校对阅读”功能。 还包括用于实施该方法的新型试剂盒。