A method or system for identification of a causative mutation causing a phenotype of interest in a test sample

Abstract

A method for identifying a mutation associated with a phenotype of interest in a non- vascular plant, wherein the method comprises (a) aligning the DNA sequence of a reference DNA sequence and identifying a first set of sequence mismatches between the two sequences; wherein the test sample is from a mutagenized non-vascular plant; (b) aligning the DNA sequence of at least one comparison sample to the reference DNA sequence and identifying a second set of sequence mismatches between the two sequences; (c) filtering the first set of mismatches with respect to the second set of mismatches to identify a subset of mismatches that are unique to the first set of mismatches, wherein the subset of mismatches are candidate mutations for the causative mutation; wherein the test sample is from a non-vascular plant exhibiting the phenotype of interest and wherein the at least one comparison sample is from an independent non- vascular plant of the same genus that does not exhibit the phenotype of interest; and wherein the reference DNA sequence is a known reference sequence for a non-vascular plant of the genus. In addition, a method for identifying a mutation associated with a phenotype of interest in a non-vascular plant, wherein the method comprises a) aligning the DNA sequence of a reference DNA sequence and identifying a first set of sequence mismatches between the two sequences; wherein the test sample is from a mutagenized non-vascular plant; (b) aligning the DNA sequence of at least one comparison sample to the reference DNA sequence and identifying a second set of sequence mismatches between the two sequences; (c) filtering the first set of mismatches with respect to the second set of sequence mismatches to identify a subset of mismatches that are common to the first and second sets of sequence mismatches wherein the test sample and the comparison sample(s) are from independent non-vascular plants exhibiting the phenotype of interest and wherein the independent non-vascular plants are the same genus; and wherein the reference DNA sequence is a known reference sequence or a non-vascular plant of the genus or a non-vascular plant of the genus.

Description

technical field

[0001] The present invention generally relates to methods or systems for identifying pathogenic mutations that cause a phenotype of interest in a test sample. Background technique

[0002] Identifying genotypes associated with phenotypes of interest is critical in many applications of plant biology. Mutation discovery following mutagenesis experiments typically involves outcrossing mutants with wild-type plants, generating large populations of wild-type and mutants, and identifying mutations that occur only in large numbers of mutants. Doing so allows genome recombination of wild-type and mutants, thus reducing the number of background mutations in the mutant genome and increasing the chances of identifying disease-causing mutations. However, this preliminary step comes at a cost.

[0003] First, the need for outcrossing represents a qualitative limitation on typical mutation discovery pipelines, since sterile mutants cannot interbreed. Many mutations (whe...

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