Detection of point mutations in the HBV polymerase gene using a fluorescence intercalator in reverse micelles

Tatsuo Maruyama, Tomoaki Takata, Hirofumi Ichinose, Noriho Kamiya, Hiroyuki Kuma, Naotaka Hamasaki, Hiroyuki Morita, Masahiro Goto

Research output: Contribution to journalArticlepeer-review

7 Citations (Scopus)

Abstract

We report a novel and simple method for mutation detection in DNA oligonucleotides using a double-stranded DNA specific dye (SYBR Green I) in nanostructured molecular assemblies, called reverse micelles. The intercalation of SYBR Green I into the duplex DNA exhibits fluorescent emission in a CTAB/isooctane reverse micellar system as well as in an aqueous solution. We found marked differences in the fluorescence intensity between perfectly matched and mismatched 52-mer synthetic oligonucleotides, which were designed to contain the YMDD motif of the hepatitis B virus (HBV) polymerase gene, in a reverse micellar solution. Using this method, we successfully detected a mutation in PCR-amplified oligonucleotides of the HBV polymerase gene in sera of four patients with chronic hepatitis B. This detection method does not require DNA immobilization, chemical modification of DNA, or any special apparatus; it only needs a normal fluorescence spectrophotometer, an inexpensive dye, and just 10 pmol of sample DNA.

Original languageEnglish
Pages (from-to)575-579
Number of pages5
JournalBiotechnology Progress
Volume21
Issue number2
DOIs
Publication statusPublished - Mar 2005

All Science Journal Classification (ASJC) codes

  • Biotechnology

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