Methods and systems for processing polynucleotides
First Claim
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1. A method for polynucleotide barcoding, comprising:
- (a) providing a first set of partitions comprising (i) a first set of nucleic acid barcode molecules comprising barcode sequences and (ii) a plurality of polynucleotides and attaching said first set of nucleic acid barcode molecules to said plurality of polynucleotides to generate a first set of barcoded polynucleotides;
(b) pooling contents of said first set of partitions;
(c) partitioning (i) a second set of nucleic acid barcode molecules comprising barcode sequences and (ii) at least a portion of said contents from (b) into a second set of partitions different than said first set of partitions; and
(d) in said second set of partitions, attaching said second set of nucleic acid barcode molecules to said first set of barcoded polynucleotides to generate a second set of barcoded polynucleotides.
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Abstract
This disclosure provides methods and compositions for sample processing, particularly for sequencing applications. Included within this disclosure are bead compositions, such as diverse libraries of beads attached to large numbers of oligonucleotides containing barcodes. Often, the beads provides herein are degradable. For example, they may contain disulfide bonds that are susceptible to reducing agents. The methods provided herein include methods of making libraries of barcoded beads as well as methods of combining the beads with a sample, such as by using a microfluidic device.
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28 Claims
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1. A method for polynucleotide barcoding, comprising:
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(a) providing a first set of partitions comprising (i) a first set of nucleic acid barcode molecules comprising barcode sequences and (ii) a plurality of polynucleotides and attaching said first set of nucleic acid barcode molecules to said plurality of polynucleotides to generate a first set of barcoded polynucleotides; (b) pooling contents of said first set of partitions; (c) partitioning (i) a second set of nucleic acid barcode molecules comprising barcode sequences and (ii) at least a portion of said contents from (b) into a second set of partitions different than said first set of partitions; and (d) in said second set of partitions, attaching said second set of nucleic acid barcode molecules to said first set of barcoded polynucleotides to generate a second set of barcoded polynucleotides. - 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)
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Specification