Methods for analyzing DNA
First Claim
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1. A method for detecting DNA in a sample, the method comprising:
- denaturing a double-stranded DNA in the sample into a first strand and a second strand;
partitioning the first strand and the second strand into a plurality of droplets, wherein each droplet contains only the first strand or only the second strand;
detecting a presence of the first strand or the second strand in each droplet; and
quantifying a number of droplets in which the first strand or the second strand is present, wherein quantifying comprises counting droplets.
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Abstract
The invention generally relates to methods for increasing the amount of DNA available for analysis when using partitioned samples and parallel processing. For example, double-stranded DNA can be dissociated into two single-stranded components, and the single strands partitioned into different droplets prior to analysis. The disclosed methods are useful for performing digital PCR analysis on samples where the target DNA is not in abundance, for example when the sample originates from a body fluid or an FFPE sample.
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Citations
14 Claims
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1. A method for detecting DNA in a sample, the method comprising:
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denaturing a double-stranded DNA in the sample into a first strand and a second strand; partitioning the first strand and the second strand into a plurality of droplets, wherein each droplet contains only the first strand or only the second strand; detecting a presence of the first strand or the second strand in each droplet; and quantifying a number of droplets in which the first strand or the second strand is present, wherein quantifying comprises counting droplets. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13)
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14. A method for determining the progression of a disease in a subject, the method comprising:
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obtaining a biological sample comprising a plurality of double-stranded DNAs from a subject; dissociating the plurality of double-stranded DNAs into a plurality of first strands and a plurality of second strands; partitioning the plurality of first strands and the plurality of second strands into a plurality of partitioned portions, at least some of the partitioned portions having only a first strand or only a second strand; detecting the presence or absence of a targeted sequence in the plurality of partitioned portions; and determining the number of occurrences of the targeted sequence by counting droplets comprising either the first strand or the second strand, wherein the number of occurrences of the targeted sequence is indicative of a progression of a disease.
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Specification