Methods and compositions for tagging and identifying polynucleotides
First Claim
1. A method of sample analysis, comprising:
- a) separately ligating an oligonucleotide tag to each of a plurality of samples that comprise fragmented genomic DNA to produce a plurality of adaptor-ligated samples, wherein;
i. each of said samples is made from a different source;
ii. each of said samples is ligated to a different oligonucleotide tag;
iii. each oligonucleotide tag comprises at least one word comprising a unique nucleotide sequence that distinguishes said oligonucleotide tag from other oligonucleotide tags; and
iv. each adaptor-ligated sample comprises adaptor-ligated genomic fragments that contain an adaptor and a genomic fragment;
b) pooling said plurality of adaptor-ligated samples to make a pooled sample;
c) isolating from said pooled sample adaptor-ligated genomic fragments that correspond to one or more genomic loci, thereby producing isolated fragments; and
d) sequencing an isolated fragment of (c) to obtain;
i. the unique nucleotide sequence of an oligonucleotide tag; and
ii. the nucleotide sequence of at least part of a genomic fragment;
wherein the unique nucleotide sequence of said isolated fragment identifies from which of said plurality of different polynucleotide samples said isolated fragment is derived.
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Abstract
The invention provides methods and compositions for attaching oligonucleotide tags to polynucleotides for the purpose of carrying out analytical assays in parallel and for decoding the oligonucleotide tags of polynucleotides selected in such assays. Words, or subunits, of oligonucleotide tags index submixtures in successively more complex sets of submixtures (referred to herein as “tiers” of submixtures) that a polynucleotide goes through while successive words are added to a growing tag. By identifying each word of an oligonucleotide tag, a series of submixtures is identified including the first submixture that contains only a single polynucleotide, thereby providing the identity of the selected polynucleotide. The analysis of the words of an oligonucleotide tag can be carried out in parallel, e.g. by specific hybridization of the oligonucleotide tag to its tag complement on an addressable array; or such analysis can be carried out serially by successive specific hybridizations of labeled word complements, or the like.
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Citations
12 Claims
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1. A method of sample analysis, comprising:
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a) separately ligating an oligonucleotide tag to each of a plurality of samples that comprise fragmented genomic DNA to produce a plurality of adaptor-ligated samples, wherein; i. each of said samples is made from a different source; ii. each of said samples is ligated to a different oligonucleotide tag; iii. each oligonucleotide tag comprises at least one word comprising a unique nucleotide sequence that distinguishes said oligonucleotide tag from other oligonucleotide tags; and iv. each adaptor-ligated sample comprises adaptor-ligated genomic fragments that contain an adaptor and a genomic fragment; b) pooling said plurality of adaptor-ligated samples to make a pooled sample; c) isolating from said pooled sample adaptor-ligated genomic fragments that correspond to one or more genomic loci, thereby producing isolated fragments; and d) sequencing an isolated fragment of (c) to obtain; i. the unique nucleotide sequence of an oligonucleotide tag; and ii. the nucleotide sequence of at least part of a genomic fragment; wherein the unique nucleotide sequence of said isolated fragment identifies from which of said plurality of different polynucleotide samples said isolated fragment is derived. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12)
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Specification