Patent ID: 6277578
Filing Date: 2001-08-21
Classification: C12Q

Abstract:
A nucleic acid assay method wherein the presence or absence of a predetermined nucleic acid target sequence is determined, and a nucleic acid target-containing sequence in a nucleic acid sample to be assayed is amplified to form an amplified nucleic acid sample prior to said determination, the improvement wherein the presence or absence of said predetermined nucleic acid target sequence is determined by steps that comprise:(A) admixing the amplified nucleic acid sample to be assayed with one or more nucleic acid probes to form a hybridization composition, wherein the 3'-terminal region of said nucleic acid probes (i) hybridizes with partial or total complementarity to at least one said predetermined nucleic acid target sequence when that sequence is present in the sample and (ii) includes an identifier nucleotide;(B) maintaining said hybridization composition for a time period sufficient to form a treated sample that may contain said amplified predetermined nucleic acid target sequence hybridized with a nucleic acid probe;(C) admixing the treated sample with a depolymerizing amount of an enzyme whose activity is to release one or more single nucleotides from the 3'-terminus of a hybridized nucleic acid probe to form a treated reaction mixture;(D) maintaining the treated reaction mixture for a time period sufficient to permit the enzyme to depolymerize hybridized nucleic acid and release identifier nucleotides therefrom; and(E) analyzing for the presence of released identifier nucleotides to obtain an analytical output, the analytical output indicating the presence or absence of at least one said nucleic acid target sequence in said nucleic acid sample.