Detection of single-base mutations by fluorogenic ribonuclease protection assay

Hirofumi Ichinose, Momoko Kitaoka, Nobuko Okamura, Tatsuo Maruyama, Noriho Kamiya, Masahiro Goto

Research output: Contribution to journalArticlepeer-review

17 Citations (Scopus)


The ribonuclease protection assay is a generally applicable technique for the detection of known mutations. We have developed a simple and rapid method for mutation detection based on the ribonuclease protection assay using fluorescently labeled oligodeoxyribonucleotide probes. The fluorogenic ribonuclease protection (FRAP) assay uses two differently labeled oligodeoxyribonucleotides, a donor probe and an acceptor probe, to obtain a fluorescence resonance energy transfer (FRET) signal. We have utilized the FRAP assay for the detection of a single-base mutation in the YMDD motif of the hepatic B virus DNA polymerase gene. The occurrence of mismatch-selective RNA cleavage was successfully discriminated by measuring the FRET signal between the donor and acceptor probes. Moreover, mutation sensing was successfully visualized by a UV transillumination. This simple and rapid mutation sensing method should facilitate a high-throughput mutation analysis.

Original languageEnglish
Pages (from-to)7047-7053
Number of pages7
JournalAnalytical Chemistry
Issue number21
Publication statusPublished - Nov 1 2005

All Science Journal Classification (ASJC) codes

  • Analytical Chemistry


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