Genotyping of polyploids
AssigneeKeygene N.V.
InventorsStefan John WHITE, René Cornelis Josephus HOGERS
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Abstract
The current invention pertains to a reliable method for determining the relative frequency of a sequence variant of interest in a nucleic acid sample derived from at least one polyploid cell, wherein the method uses a UMI to correct for any amplification biases. The invention further pertains to the use of a UMI for accurately determining the relative frequency of a sequence variant of interest in a nucleic acid sample derived from at least one polyploid cell.
Claims20
A method for determining the relative frequency of a sequence variant of interest in a nucleic acid sample derived from at least one polyploid cell, the method comprising:
- (a) providing polynucleotides derived from the nucleic acid sample, wherein each polynucleotide comprises a unique molecular index (UMI),
- (b) amplifying the polynucleotides provided in (a);
- (c) determining the sequences of the amplified polynucleotides to obtain sequence reads;
- (d) obtaining a consensus sequence of the sequence reads that are derived from a single polynucleotide of (a) using at least the UMI;
- and
- (e) determining the relative frequency of the sequence variant of interest based on the frequency of consensus sequences comprising the sequence variant of interest and the frequency of a reference sequence.
An allele-specific oligonucleotide probe for use in an oligonucleotide ligation assay, wherein the oligonucleotide probe comprises a UMI, comprised in at least one of:
- (i) an oligonucleotide ligation probe;
- and
- (ii) an adapter.
A kit of parts for determining the relative frequency of a sequence variant of interest in a nucleic acid sample derived from at least one polyploid cell, the kit comprising at least one of:
- (i) a vial comprising a mixture of oligonucleotide ligation probes, wherein at least part of the oligonucleotide ligation probes comprise a UMI;
- (ii) a vial comprising a mixture of adapter molecules, wherein the adapter molecules comprise a UMI and optionally a sample identifier;
- and
- (iii) a vial comprising one or more amplification primers.
Dependencies are inferred from the claim text.