US 2001/0034029 A1Published application

Method of detecting mutation in base sequence of nucleic acid

AssigneeShimadzu Corporation

InventorHideshi Fujiwake

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Abstract

Normal DNA (15) and mutational DNA (17) are mixed and subjected to PCR amplification (A). Fluorescent oligonucleotides (19a, 19b, 19c) complementary to any of exons (15a, 15b, 15c) and labeled with fluorescent materials (F1, F2, F3) having different fluorescence spectral characteristics are prepared (B) and hybridized with the amplified substance for forming homoduplexes (21a, 21b, 21c, 23b) and heteroduplexes (23a, 23c) (C). Since the heteroduplexes (23a, 23c) have a lower melting temperature than the homoduplexes (21a, 21b, 21c, 23b), analysis is made with an ion pair chromatograph having a reversed phase column set at the melting temperature for discriminating and detecting relatively quickly eluting fluorescent oligonucleotides (19a, 19c) thereby detecting mutational exons.

Claims6

Dependent claims

A method of detecting mutation in the base sequence of nucleic acid, including:

(A) a bonding step of hybridizing an object of analysis consisting of nucleic acid or a nucleic acid fragment including a plurality of inspected sites to be subjected to inspection of mutation in the base sequence with a plurality of types of oligonucleotides having base sequence complementary to any of the inspected sites having normal base sequence and labeled to be discriminable from each other for forming duplexes;
and
(B) a detection step of employing an ion pair chromatograph comprising a reversed phase column serving as a separation column and a detector capable of discriminating and detecting the labeled oligonucleotides and setting the separation column at a temperature causing difference in stability between hetero- and homoduplexes included in the duplexes for analyzing the object of analysis.

Dependencies are inferred from the claim text.