Patent ID: 7851159
Filing Date: 2010-12-14
Classification: C12Q,Y10T

Abstract:
1. A method for detecting a target nucleic acid having a specific base sequence contained in a nucleic acid mixture, wherein: utilizing a set of nucleic acids for detecting a target nucleic acid having a specific base sequence, which the set of nucleic acids includes a photoligating nucleic acid comprising a nucleic acid having a group represehted by the following Formula I, Formula II, Formula III, Formula IV or Formula V: wherein Z represents O or NH; at least one of X and Y represents an electron -withdrawing group selected from the group consisting of a carboxyl group, a lower alkoxycarbonyl group, a substituted amide group and a cyano group; and the remainder of X and Y represents a hydrogen atom; wherein R 1 is a hydrogen atom; at least one of R 2 and R 3 represents a group selected from the group consisting of a carboxyl group, a lower alkoxycarbonyl group, a lower alkenyl group, a lower alkynyl group, a substituted amide group, an amide group, a cyano group and a hydrogen atom; and the remainder of R 2 and R 3 represents a hydrogen atom or a cyano group; wherein R 4 is a hydrogen atom or a lower alkyl group; at least one of R 5 and R 6 represents a group selected from the group consisting of a carboxyl group, a lower alkoxycarbonyl group, a lower alkenyl group, a lower alkynyl group, a substituted amide group, an amide group, a cyano group and a hydrogen atom; and the remainder of R 5 and R 6 represents a hydrogen atom or a cyano group; wherein R 7 is a hydrogen atom or a lower alkyl group; at least one of R 8 and R 9 represents a group selected from the group consisting of a carboxyl group, a lower alkoxycarbonyl group, a lower alkenyl group, a lower alkynyl group, a substituted amide group, an amide group, a cyano group and a hydrogen atom; and the remainder of R 8 and R 9 represents a hydrogen atom or a cyano group; wherein R 10 is a hydrogen atom or a lower alkyl group; at least one of R 11 and R 12 represents a group selected from the group consisting of a carboxyl group, a lower alkoxycarbonyl group, a lower alkenyl group, a lower alkynyl group, a substituted amide group, an amide group, a cyano group and a hydrogen atom; and the remainder of R 11 and R 12 represents a hydrogen atom or a cyano group; at the 5′-end or 3′-end as a base moiety, and a nucleic acid to be photoligated having a base having a carbon-carbon double bond at the 3′-end or 5′-end as a base moiety capable of being photoligated to the photoligating nucleic acid, wherein any one of the photoligating nucleic acid and the nucleic acid to be photoligated has a labeling moiety, while the other one not having a labeling moiety between the photoligating nucleic acid and the nucleic acid to be photoligated is immobilized on a substrate in advance; a nucleic acid having a base sequence that is complementary to the base sequence of the target nucleic acid having a specific base sequence, can be generated by photoligation of the photoligating nucleic acid and the nucleic acid to be photoligated; and the nucleic acid that can be generated by photoligation of the photoligating nucleic acid and the nucleic acid to be photoligated, can be washed under a washing condition capable of dissociating complementary double strands, wherein the method comprises the following steps of: hybridizing the target nucleic acid having a specific base sequence contained in a nucleic acid mixture with the photoligating nucleic acid and the nucleic acid to be photoligated; immobilizing a nucleic acid having a labeling moiety on a substrate by photoirradiating the nucleic acid to result in photoligation of the nucleic acid with the photoligating nucleic acid and nucleic acid to be photoliqated that have been brought adjacent in a manner capable of photoligating through the hybridization using the target nucleic acid having a specific base sequence as the template; removing any nucleic acid having a labeling moiety and not immobilized on the substrate through photoligation by washing the nucleic acid having a labeling moiety and immobilized on the substrate through photoligation, under a washing condition capable of dissociating hybridized complementary double strands; and detecting the labeling moiety so as to detect the nucleic acid having a labeling moiety and immobilized on the substrate through photoligation; wherein the washing condition involves washing at a washing temperature in the range of 80° C. to 100° C.; wherein the process of hybridization and the process of immobilizing a nucleic acid having a labeling moiety through photoligation on the substrate are carried out in a reaction solution containing a salt having buffering effects; wherein the reaction solution contains 20% to 60% v/v of a water-dispersible organic solvent; and wherein the salt having buffering effects is a cacodylic acid salt.