Method for determining the presence of an RNA analyte in a sample using a modified oligonucleotide probe
First Claim
1. A diagnostic method for the specific detection of an RNA analyte in a sample, said method comprising:
- a) processing a sample to allow for or enhance discrimination between an RNA analyte and non-analyte nucleic acids present in said sample;
b) contacting said sample with an oligonucleotide probe having a detectable label under conditions such that a first base region of said probe stably and specifically hybridizes to a second base region of said RNA analyte to form a probe;
analyte hybrid containing at least one ribonucleotide modified to include a 2′
-O-methyl substitution to the ribofuranosyl moiety, wherein said probe does not stably hybridize under said conditions to non-analyte nucleic acids present in said sample, and wherein said probe is free in solution when contacted with said sample; and
c) determining whether said hybrid is present in said sample as an indication of the presence or absence of said RNA analyte in said sample.
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Accused Products
Abstract
The present invention concerns oligonucleotides containing one or more modified nucleotides which increase the binding affinity of the oligonucleotides to target nucleic acids having a complementary nucleotide base sequence. These modified oligonucleotides hybridize to the target sequence at a faster rate than unmodified oligonucleotides having an identical nucleotide base sequence. Such modified oligonucleotides include oligonucleotides containing at least one 2′-O-methylribofuranosyl moiety joined to a nitrogenous base. Oligonucleotides can be modified in accordance with the present invention to preferentially bind RNA targets. The present invention also concerns methods of using these modified oligonucleotides and kits containing the same.
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Citations
109 Claims
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1. A diagnostic method for the specific detection of an RNA analyte in a sample, said method comprising:
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a) processing a sample to allow for or enhance discrimination between an RNA analyte and non-analyte nucleic acids present in said sample;
b) contacting said sample with an oligonucleotide probe having a detectable label under conditions such that a first base region of said probe stably and specifically hybridizes to a second base region of said RNA analyte to form a probe;
analyte hybrid containing at least one ribonucleotide modified to include a 2′
-O-methyl substitution to the ribofuranosyl moiety, wherein said probe does not stably hybridize under said conditions to non-analyte nucleic acids present in said sample, and wherein said probe is free in solution when contacted with said sample; and
c) determining whether said hybrid is present in said sample as an indication of the presence or absence of said RNA analyte in said sample. - 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, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 107)
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38. A diagnostic method for the specific detection of an RNA analyte in a sample, said method comprising:
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a) contacting a sample with an oligonucletide probe having a detectable label in the presence of a detergent under conditions such that a first base region of said probe stably and specifically hybridizes to a second base region of an RNA analyte to form a probe;
analyte hybrid containing at least one ribonucleotide modified to include a 2′
-O-methyl substitution to the ribofuranosyl moiety, wherein said probe does not stably hybridize under said conditions to non-analyte nucleic acids present in said sample; and
b) determining whether said hybrid is present in said sample as an indication of the presence or absence of said RNA analyte in said sample. - View Dependent Claims (39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 108)
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73. A diagnostic method for the specific detection of an RNA analyte in a sample, said method comprising:
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a) contacting a sample with an oligonucleotide probe having a detectable label under conditions such that a first base region of said probe stably and specifically hybridizes to a second base region present in an RNA analyte to form a probe;
analyte hybrid containing at least one ribonucleotide modified to include a 240 -O-methyl substitution to the ribofuranosyl moiety, said first base region targeting a double-stranded region contained within said second base region, wherein said probe does not stably hybridize under said conditions to non-analyte nucleic acids present in said sample, and wherein said probe is free in solution when contacted with said sample; and
b) determining whether said hybrid is present in said sample as an indication of the presence or absence of said RNA analyte in said sample. - View Dependent Claims (74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 109)
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Specification