Systems and methods to detect rare mutations and copy number variation

  • US 10,704,086 B2
  • Filed: 10/04/2019
  • Issued: 07/07/2020
  • Est. Priority Date: 03/05/2014
  • Status: Active Grant
First Claim
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1. A method for detecting a presence or absence of one or more somatic genetic variants in cell-free deoxyribonucleic acid (cfDNA) molecules from a bodily fluid sample of a subject, comprising:

  • (a) non-uniquely tagging a plurality of cfDNA molecules from a population of cfDNA molecules obtained from the bodily fluid sample with molecular barcodes from a set of molecular barcodes to produce non-uniquely tagged parent polynucleotides,wherein the non-uniquely tagging comprises ligating molecular barcodes from the set of molecular barcodes to both ends of a cfDNA molecule from the plurality of cfDNA molecules using more than a 10×

    molar excess of molecular barcodes relative to the population of cfDNA molecules,wherein the cfDNA molecules that map to a mappable base position of a reference sequence are tagged with a number of diffrent molecular barcodes ranging from at least 2 and fewer than a number of cfDNA molecules that map to the mappable base position, andwherein at least 20% of the cfDNA molecules from the population of cfDNA molecules are attached to molecular barcodes;

    (b) amplifying a plurality of the non-uniquely tagged parent polynucleotides to produce progeny polynucleotides with associated molecular barcodes;

    (c) sequencing a plurality of the progeny polynucleotides to produce sequencing reads of the progeny polynucleotides with associated molecular barcodes;

    (d) mapping a plurality of the sequencing reads to the reference sequence to generate mapped sequencing reads;

    (e) grouping a plurality of the mapped sequencing reads into a plurality of families based on sequence information from the molecular barcodes and at least (1) a start base position of a given mapped sequencing read from among the mapped sequencing reads at which the given mapped sequencing read is determined to start mapping to the reference sequence and/or (2) a stop base position of the given mapped sequencing read at which the given mapped sequencing read is determined to stop mapping to the reference sequence; and

    (f) detecting, from among the mapped sequencing reads in a plurality of the families, the presence or absence of the one or more somatic genetic variants.

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