Expression monitoring by hybridization to high density oligonucleotide arrays
First Claim
1. A method of analyzing the expression of one or more genes, said method comprising:
- (a) providing a pool of target nucleic acids comprising RNA transcripts of one or more of said genes, or nucleic acids derived therefrom using said RNA transcripts as templates;
(b) hybridizing said pool of tarzet nucleic acids to an array of oligonucleotide probes immobilized on a surface, said array comprising more than 100 different oligonucleotides, at least some of which comprise control probes, wherein each different oligonucleotide is localized in a predetermined region of said surface, the density of said different oligonucleotides is greater than about 60 different oligonucleotides per 1 cm2, and at least some of said oligonucleotide probes are complementary to said RNA transcripts or said nucleic acids derived therefrom using said RNA transcripts; and
(c) quantifying the hybridization of said nucleic acids to said array, wherein said quantification is proportional to the expression level of said genes.
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Abstract
This invention provides methods of monitoring the expression levels of a multiplicity of genes. The methods involve hybridizing a nucleic acid sample to a high density array of oligonucleotide probes where the high density array contains oligonucleotide probes complementary to subsequences of target nucleic acids in the nucleic acid sample. In one embodiment, the method involves providing a pool of target nucleic acids comprising RNA transcripts of one or more target genes, or nucleic acids derived from the RNA transcripts, hybridizing said pool of nucleic acids to an array of oligonucleotide probes immobilized on surface, where the array comprising more than 100 different oligonucleotides and each different oligonucleotide is localized in a predetermined region of the surface, the density of the different oligonucleotides is greater than about 60 different oligonucleotides per 1 cm2, and the olignucleotide probes are complementary to the RNA transcripts or nucleic acids derived from the RNA transcripts; and quantifying the hybridized nucleic acids in the array.
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Citations
45 Claims
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1. A method of analyzing the expression of one or more genes, said method comprising:
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(a) providing a pool of target nucleic acids comprising RNA transcripts of one or more of said genes, or nucleic acids derived therefrom using said RNA transcripts as templates; (b) hybridizing said pool of tarzet nucleic acids to an array of oligonucleotide probes immobilized on a surface, said array comprising more than 100 different oligonucleotides, at least some of which comprise control probes, wherein each different oligonucleotide is localized in a predetermined region of said surface, the density of said different oligonucleotides is greater than about 60 different oligonucleotides per 1 cm2, and at least some of said oligonucleotide probes are complementary to said RNA transcripts or said nucleic acids derived therefrom using said RNA transcripts; and (c) quantifying the hybridization of said nucleic acids to said array, wherein said quantification is proportional to the expression level of said genes. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 37, 38, 41, 42, 43, 44, 45)
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24. A method of identifying oligonucleotide probes to quantify the expression of a target nucleic acid, comprising:
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providing a high density array of oligonucleotide probes, said array comprising a multiplicity of oligonucleotide probes, wherein each probe is complementary to a subsequence of said target nucleic acid and for each probe there is a corresponding mismatch control probe; hybridizing said target nucleic acid to said array of oligonucleotide probes; and identifying those probes for which the difference in hybridization signal intensity between each probe and its mismatch control is detectable. - View Dependent Claims (25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36)
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39. A method of identifg polynucleotide probes to quantify the expression of a target nucleic acid, comprising:
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providing a high density array comprising polynucleotide probes complementary to a subsequence of said target nucleic acid, wherein for each of said probes there is a corresponding mismatch control probe on said array; hybridizing said array to a pool of nucleic acids comprising nucleic acids other than said target nucleic acids; and identifying probes having a hybridization intensity equal to or less than a background intensity, wherein at least one of said probes chosen is used to quantify said expression of said target nucleic acid. - View Dependent Claims (40)
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Specification