Method for Detection of Single-Base Mismatches Using Bimolecular Beacons
- 26 February 2002
- journal article
- research article
- Published by American Chemical Society (ACS) in Journal of the American Chemical Society
- Vol. 124 (11), 2396-2397
- https://doi.org/10.1021/ja012374d
Abstract
This paper describes a method for the detection of single-base mismatches using DNA microarrays in a format that does not require labeling of the sample (“target”) DNA. The method is based on disrupting fluorescence energy transfer (FRET) between a fluorophore attached to an immobilized DNA strand (“probe”) and a quencher-containing sequence that is complementary except for an artificial mismatch (e.g. 5-nitroindole, 3-nitropyrole, or abasic site) at the site of interrogation. As the displacement of the FRET acceptor and hybridization of the unlabeled probe are bimolecular, the term “bimolecular beacons” is used to describe this approach. The analysis of a mismatch was based on differences in the amount of disruption in FRET upon hybridization of perfectly matched DNA targets and those containing single-base mismatches. Using this method and an oligonucleotide model system, A/C single-base mismatches were successfully discriminated at levels greater than that observed using surface-immobilized molecular beacons. The amount of discrimination was dependent on the identity of the artificial mismatch; greater discrimination was observed with 5-nitroindole (a “universal” base) than with an abasic site. G/T mismatches, considered to be particularly difficult to detect, were also successfully discriminated when quencher sequences containing 5-nitroindole were used.Keywords
This publication has 15 references indexed in Scilit:
- Single-nucleotide polymorphism analysis by MALDI–TOF mass spectrometryTrends in Biotechnology, 2000
- The essence of SNPsGene, 1999
- Resequencing and mutational analysis using oligonucleotide microarraysNature Genetics, 1999
- Exploring the new world of the genome with DNA microarraysNature Genetics, 1999
- Shotgun Sequencing of the Human GenomeScience, 1998
- RasGRP, a Ras Guanyl Nucleotide- Releasing Protein with Calcium- and Diacylglycerol-Binding MotifsScience, 1998
- Multicolor molecular beacons for allele discriminationNature Biotechnology, 1998
- Energy-transfer fluorescent reagents for DNA analysesCurrent Opinion in Biotechnology, 1997
- Solution-phase detection of polynucleotides using interacting fluorescent labels and competitive hybridizationAnalytical Biochemistry, 1989
- Dissociation kinetics of 19 base paired oligonucleotide-DNA duplexes containing different single mismatched base pairsNucleic Acids Research, 1987