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Real-time PCR method for detection of short DNA using a deoxyuridine probe and application for detection of
Naoki Harikai1, Yuko Tanaka1, Satoshi Miyashita1
1School of Pharmacy, Nihon University, 7-7-1 Narashinodai, Funabashi, Chiba, 274-8555, Japan.
Abstract:
This study sought to develop a short DNA detection method using a deoxyuridine probe and polymerase chain reaction. The probe was hybridized to the target short DNA, which was then extended by DNA polymerase. The extended DNA was used for real-time PCR after the probe was removed by uracil DNA glycosylase. This method measured from 0.01 to 10 nM of a model short DNA sequence of 17 nucleotides. The method was then used to detect the nucleic acid medicine fomivirsen, as well as 21 phosphorothioate nucleotides, and to quantify 0.1-100 nM of fomivirsen. This method may be useful for detecting short DNA fragments, such as functional nucleotides.
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