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 journalArticle

16 Citations (Scopus)

Abstract

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
Volume77
Issue number21
DOIs
Publication statusPublished - Nov 1 2005

All Science Journal Classification (ASJC) codes

  • Analytical Chemistry

Fingerprint Dive into the research topics of 'Detection of single-base mutations by fluorogenic ribonuclease protection assay'. Together they form a unique fingerprint.

  • Cite this