Tag reagent and assay method
First Claim
1. A method of sequencing a target nucleic acid, which method comprises the steps of:
- a) providing an oligonucleotide immobilized on a support,b) hybridizing the target nucleic acid with the immobilized oligonucleotide,c) incubating the hybrid from b) with a library comprising a solution of reagents, each of said reagents comprising1) an analyte moiety comprising at least two analyte residues, and linked to2) a tag moiety comprising one or more reporter groups suitable for detection by mass spectrometry, excluding oligonucleotides.wherein a reporter group designates an analyte residue, and the reporter group at each position of the tag moiety is chosen to designate an analyte residue at a defined position of the analyte moietyand wherein the library consists of 4n reagents, each of said reagents comprising a different analyte moiety which is a different oligonucleotide chain of n nucleotides,so that an oligonucleotide chain of a first reagent of the library becomes hybridized to the target nucleic acid adjacent the immobilized oligonucleotide,d) ligating the adjacent oligonucleotides, thus forming a ligated first reagent,e) removing other non-ligated reagents, andf) recovering and analyzing the tag moiety of the ligated first reagent as an indication of the sequence of a first part of the target nucleic acid wherein the said target nucleic acid and ligated first reagent remain hybridized.
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Accused Products
Abstract
A reagent comprises: a) an analyte moiety comprising at least two analyte residues, and linked to; b) a tag moiety comprising one or more reporter groups adapted for detection by mass spectrometry, wherein a reporter group designates analyte residue, and the reporter group at each position of the tag moiety is chosen to designate an analyte residue at a defined position of the analyte moiety. A plurality of such reagents, each comprising a different analyte moiety, provides a library of reagents which may be used in assay methods involving a target substance. Analysis of the tag moieties indicates the nature of the analyte moieties bound to the target substance. A method of sequencing nucleic acid employs a library of the reagents to determine the sequence of a target nucleic acid.
394 Citations
10 Claims
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1. A method of sequencing a target nucleic acid, which method comprises the steps of:
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a) providing an oligonucleotide immobilized on a support, b) hybridizing the target nucleic acid with the immobilized oligonucleotide, c) incubating the hybrid from b) with a library comprising a solution of reagents, each of said reagents comprising 1) an analyte moiety comprising at least two analyte residues, and linked to 2) a tag moiety comprising one or more reporter groups suitable for detection by mass spectrometry, excluding oligonucleotides. wherein a reporter group designates an analyte residue, and the reporter group at each position of the tag moiety is chosen to designate an analyte residue at a defined position of the analyte moiety and wherein the library consists of 4n reagents, each of said reagents comprising a different analyte moiety which is a different oligonucleotide chain of n nucleotides, so that an oligonucleotide chain of a first reagent of the library becomes hybridized to the target nucleic acid adjacent the immobilized oligonucleotide, d) ligating the adjacent oligonucleotides, thus forming a ligated first reagent, e) removing other non-ligated reagents, and f) recovering and analyzing the tag moiety of the ligated first reagent as an indication of the sequence of a first part of the target nucleic acid wherein the said target nucleic acid and ligated first reagent remain hybridized. - View Dependent Claims (2, 3, 4, 5, 6)
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7. A library of reagents, wherein each reagent comprises
a) an analyte moiety comprising at least two analyte residues, and linked to b) a tag moiety comprising one or more reporter groups suitable for detection by mass spectrometry, excluding oligonucleotides, wherein a reporter group designates an analyte residue, and the reporter group at each position of the tag moiety is chosen to designate an analyte residue at a defined position of the analyte moiety, wherein the library consists of 4n reagents each comprising a different analyte moiety which is a different oligonucleotide chain of n nucleotides.
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9. A method of analyzing a target DNA, which method comprises the steps of:
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i) providing the target DNA immobilized on a support, ii) incubating the immobilized target DNA from i) with a plurality of reagents where each reagent comprises a) an analyte moiety which is an oligonucleotide chain comprising at least two nucleotide residues, and linked to b) a tag moiety comprising one or more reporter groups suitable for detection by mass spectrometry, excluding oligonucleotides, wherein a reporter group designates an analyte residue, and the reporter group at each position of the tag moiety is chosen to designate an analyte residue at a defined position of the analyte moiety, so that the oligonucleotide chains of different reagents become hybridized to the target DNA on the support, iii) removing non-hybridized reagents, and iv) recovering and analyzing the tag moiety of each reagent as an indication of the sequence of a part of the target DNA wherein the said target DNA and any hybridized reagents remain hybridized. - View Dependent Claims (10)
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Specification