Sensitive mutant detection by concentrating mutant DNA with allele-specific capture and its application to analysis of contaminated grains in rice

Ryuya Kohata, Kosuke Koitabashi, Hiroyasu Kitashiba, Takeshi Nishio

    Research output: Contribution to journalArticlepeer-review

    3 Citations (Scopus)

    Abstract

    Key message: We developed a method for detection of mutants in a large number of plants, and found this method to be applicable to detection of a mutant allele at a concentration of 1/1000. Abstract: Many techniques for SNP analysis have been developed, but most of these techniques are not so sensitive to be used for detection of mutants in a large number of plants. Although some highly sensitive methods of SNP analysis have been reported, they are costly. In the present study, a method for concentrating mutant DNA was examined for sensitive detection of an SNP allele in a bulked DNA sample. PCR products of mutant alleles were captured by biotin-labeled oligonucleotide conjugated with streptavidin-coated magnetic beads. By repeated captures of each strand and combining both strands, mutant alleles with a concentration of 1/1000 in wild-type alleles were detectable by CAPS or dCAPS analysis. Indirect capture of a mutant allele was possible, but efficiency was slightly lower than that of the direct capture. The developed method was applied to detection of contamination of rice grains by grains of a different cultivar. Possible applications of this method are discussed.

    Original languageEnglish
    Pages (from-to)865-872
    Number of pages8
    JournalPlant Cell Reports
    Volume37
    Issue number6
    DOIs
    Publication statusPublished - 2018 Jun 1

    Keywords

    • Cost-effective method
    • High sensitivity
    • Mutation detection
    • SNP analysis
    • Seed contamination test

    ASJC Scopus subject areas

    • Agronomy and Crop Science
    • Plant Science

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