METHODS AND MATERIALS FOR ASSESSING RNA EXPRESSION

摘要

This document provides methods and materials for assessing RNA expression. For example, methods and materials for detecting the presence, absence, or amount of target nucleic acid (e.g., target RNA or target cDNA produced from target RNA), kits for detecting the presence, absence, or amount of target nucleic acid (e.g., target RNA or target cDNA produced from target RNA), and methods for making such kits are provided.

主权项

1. A method for assessing RNA expression, said method comprising:
(a) contacting a sample to be assessed for expression of a target RNA with a probe nucleic acid comprising an amplifying restriction endonuclease and a nucleotide sequence complementary to a sequence of said target RNA or a cDNA of said target RNA under conditions wherein, if said target RNA or said cDNA is present in said sample, at least a portion of said target RNA or said cDNA hybridizes to at least a portion of said probe nucleic acid to form a double-stranded portion of nucleic acid comprising a restriction endonuclease cut site, (b) contacting said double-stranded portion of nucleic acid with a recognition restriction endonuclease having the ability to cut said double-stranded portion of nucleic acid at said restriction endonuclease cut site under conditions wherein said recognition restriction endonuclease cleaves said double-stranded portion of nucleic acid at said restriction endonuclease cut site, thereby separating a portion of said probe nucleic acid comprising said amplifying restriction endonuclease from at least another portion of said probe nucleic acid, (c) contacting said portion of said probe nucleic acid comprising said amplifying restriction endonuclease with a reporter nucleic acid comprising a double-stranded portion of nucleic acid comprising a restriction endonuclease cut site of said amplifying restriction endonuclease under conditions wherein said amplifying restriction endonuclease cleaves said reporter nucleic acid at said restriction endonuclease cut site of said amplifying restriction endonuclease, thereby separating a portion of said reporter nucleic acid from at least another portion of said reporter nucleic acid, and (d) determining the presence or absence of said portion of said reporter nucleic acid, wherein the presence of said portion of said reporter nucleic acid indicates that said sample contains said target RNA or said cDNA, thereby indicating expression of said target RNA, and wherein the absence of said portion of said reporter nucleic acid indicates that said sample does not contain said target RNA or said cDNA, thereby indicating a lack of expression of said target RNA.