METHODS AND COMPOSITIONS FOR DETECTING TARGET NUCLEIC ACIDS

摘要

The present invention provides compositions, apparatuses and methods for detecting one or more nucleic acid targets present in a sample. Methods of the invention include utilizing two or more ligation probes that reversibly bind a target nucleic acid in close proximity to each other and possess complementary reactive ligation moieties. When such probes have bound to the target in the proper orientation, they are able to undergo a spontaneous chemical ligation reaction that yields a ligation product that is directly detected or that is amplified to produce amplicons that are then detected. The present invention also provides methods to stabilize sample RNA so that degradation does not significantly affect the results of the analysis.

主权项

1. A method for detecting a plurality of different target nucleic acids in a sample, wherein each target nucleic acid comprises a first target domain adjacent to a second target domain and a third target capture domain located upstream or downstream from said first and second target domains, said method comprising:
(a) providing a plurality of ligation substrates each comprising:
(i) one of said target nucleic acids;(ii) a first set of ligation probes comprising:
(A) a first nucleic acid ligation probe comprising:
(1) a first probe domain hybridized to a first target domain of said one target nucleic acid sequence;(2) a 5′-ligation moiety; and(B) a second nucleic acid ligation probe comprising:
(1) a second probe domain hybridized to a second target domain of said one target nucleic acid sequence; and(2) a 3′ ligation moiety; (b) ligating said first and second ligation probes without the use of a ligase enzyme to form a first plurality of ligation products; (c) hybridizing target capture probes comprising a capture moiety to said third target domain of said target nucleic acids to form target complexes; (d) capturing said target complexes on a surface using said capture moiety; (e) amplifying said ligation products to form amplicons; (f) detecting said amplicons, thereby detecting said target nucleic acids.