| 摘要 |
The detection of variant nucleotides in a nucleic acid template may be achieved by contacting the template with a first oligonucleotide, a second oligonucleotide and a third oligonucleotide. The nucleic acid template has first, second and third regions. The first region is adjacent to and in the 3' direction from the second region, and the first oligonucleotide is substantially complementary to the first region and has a 3' end that is not extendible by a polymerase; the second region is between and adjacent to the first and third regions, and contains a site to be tested for the presence or absence of a variant nucleotide, the second region and second oligonucleotide having an identical number of nucleotides, the second oligonucleotide having a 3' end that is not extendible by a polymerase, and a nucleotide sequence such that (a) the second oligonucleotide hybridizes to the second region when the variant nucleotide in the second region is complementary to the corresponding nucleotide in the second oligonucleotide; and (b) the second oligonucleotide does not hybridize to the second region when the variant nucleotide in the second region is not complementary to the corresponding nucleotide in the second oligonucleotide; and the third region is adjacent to and in the 5' direction from the second region, the third oligonucleotide being substantially complementary to the third region and having a 3' end that is extendible by a polymerase. The template nucleic acid is contacted with the three oligonucleotides under reaction conditions where the 3' end of the third oligonucleotide undergoes an extension by polymerase depending on whether the second region contains the variant nucleotide.
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