Nucleic acid indexing techniques
AssigneeIllumina, Inc.
InventorsJohn S. Vieceli, Ryan Matthew Kelley
Life of this patent
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Abstract
Presented herein are techniques for indexing of nucleic acid, e.g., for use in conjunction with sequencing. The techniques include generating indexed nucleic acid fragments from an individual sample, whereby the index sequence incorporated into each index site of the nucleic acid fragment is selected from a plurality of distinguishable of index sequences and such that the population of generated nucleic acid fragments represents each index sequence from the plurality. In this manner, the generated indexed nucleic acid fragments from a single sample are indexed with a diverse mix of index sequences that reduce misassignment due to index read errors associated with low sequence diversity.
Claims10
A method for sequencing nucleic acid molecules, comprising:
- providing a plurality of dual-indexed nucleic acid fragments generated from a sample, wherein each individual nucleic acid fragment of the plurality of dual-indexed nucleic acid fragments comprises a 5′ adapter sequence, a 5′ index sequence, a target sequence, a 3′ adapter sequence, and a 3′ index sequence, wherein a plurality of different 5′ index sequences selected from a first set of 5′ index sequences associated with the sample and a plurality of different 3′ index sequences selected from a second set of 3′ index sequences associated with the sample are represented in the plurality of dual-indexed nucleic acid fragments, wherein the plurality of different 5′ index sequences and the plurality of different 3′ index sequences are distinguishable from one another, and wherein the plurality of dual-indexed nucleic acid fragments comprises more than one combination of the 5′ index sequence and the 3′ index sequence such that one or more individual fragments of the plurality of dual-indexed nucleic acid fragments have respective different 5′ index sequences and respective different 3′ index sequences relative to one another, wherein more than one fragment of the plurality has a same combination of an index sequence of the first set and an index sequence of the second set but different target sequences relative to one another;
- generating sequencing data representative of sequences of the plurality of dual-indexed nucleic acid fragments;
- and
- associating an individual sequence of the sequences with the sample only when the individual sequence includes both the 5′ index sequence selected from the first set and the 3′ index sequence selected from the second set.
A method for sequencing nucleic acid molecules, comprising:
- providing a plurality of dual-indexed nucleic acid fragments generated from a sample, wherein each individual nucleic acid fragment of the plurality of dual-indexed nucleic acid fragments comprises a sequence of interest derived from the sample, a 5′ adapter sequence, a 5′ index sequence, a 3′ adapter sequence, and a 3′ index, wherein a plurality of different 5′ index sequences selected from a first set of 5′ index sequences associated with the sample and a plurality of different 3′ index sequences selected from a second set of 3′ index sequences associated with the sample are represented in the plurality of dual-indexed nucleic acid fragments and wherein the plurality of different 5′ index sequences and the plurality of different 3′ index sequences are distinguishable from one another and wherein the plurality of dual-indexed nucleic acid fragments comprises more than one combination of the 5′ index sequence and the 3′ index sequence such that one or more individual fragments of the plurality of dual-indexed nucleic acid fragments have respective different 5′ index sequences and respective different 3′ index sequences relative to one another, wherein more than one fragment of the plurality has a same combination of an index sequence of the first set and an index sequence of the second set but different sequences of interest relative to one another;
- generating sequencing data representative of the sequence of interest;
- generating sequencing data representative of the 5′ index sequence and the 3′ index sequence;
- and
- assigning an individual sequence of interest to the sample only when the individual sequence of interest is associated with both the 5′ index sequence selected from the first set and the 3′ index sequence selected from the second set.
Dependencies are inferred from the claim text.