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Curr Protoc Mol Biol. 2014 Oct 1;108:7.18.1-16. doi: 10.1002/0471142727.mb0718s108.

Next-Gen Sequencing-Based Mapping and Identification of Ethyl Methanesulfonate-Induced Mutations in Arabidopsis thaliana.

Author information

1
Department of Genetics, Harvard Medical School and Department of Molecular Biology, Massachusetts General Hospital, Boston, Massachusetts.

Abstract

Forward genetic analysis using ethyl methanesulfonate (EMS) mutagenesis has proven to be a powerful tool in biological research, but identification and cloning of causal mutations by conventional genetic mapping approaches is a painstaking process. Recent advances in next-gen sequencing have greatly invigorated the process of identifying EMS-induced mutations corresponding to a specific phenotype in model genetic hosts, including the plant Arabidopsis thaliana and the nematode Caenorhabditis elegans. Next-gen sequencing of bulked F2 mutant recombinants produces a wealth of high-resolution genetic data, provides enhanced delimitation of the genomic location of mutations, and greatly reduces hands-on time while maintaining high accuracy and reproducibility. In this unit, a detailed procedure to simultaneously map and identify EMS mutations in Arabidopsis is described.

KEYWORDS:

ethyl methanesulfonate (EMS); genetic mapping; next-gen sequencing

PMID:
25271717
DOI:
10.1002/0471142727.mb0718s108
[Indexed for MEDLINE]

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