Patent ID: 7485425

Claim:
A method for determining whether at least one target nucleic acid sequence is present in a sample, comprising: forming a ligation reaction composition comprising the sample, and a ligation probe set for each target nucleic acid sequence, the probe set comprising (a) a first probe, comprising a first target-specific portion, and (b) a second probe, comprising a second target-specific portion, wherein the probes in each set are suitable for ligation together when hybridized adjacent to one another on the complementary target nucleic acid sequence; forming a first test composition by subjecting the ligation reaction composition to at least one cycle of ligation, wherein adjacently hybridizing complementary probes are ligated to one another to form a ligation product comprising the first probe and the second probe if a target nucleic acid sequence is present in the sample; forming an amplification reaction composition comprising: at least some of the first test composition; a polymerase; a first primer comprising (i) a sequence complementary to the 5′ end of the ligation product and (ii) a sequence complementary to the 3′ end of the ligation product; forming a second test composition by subjecting the amplification reaction composition to at least one amplification reaction, wherein the second test composition comprises at least one amplification product if a target nucleic acid sequence is present in the sample; and wherein the 5′ end of the ligation product and the 3′ end of the ligation product are not ligated to one another; and determining whether the at least one target nucleic acid sequence is present by detecting at least one amplification product.