title: Next-Gen Sequencing-Based Mapping and Identification of Ethyl Methanesulfonate-Induced Mutations in Arabidopsis thaliana.
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.
authors.affiliation: Department of Genetics, Harvard Medical School and Department of Molecular Biology, Massachusetts General Hospital, Boston, Massachusetts.
authors.forename: Xue-Cheng
authors.initials: XC
authors.lastname: Zhang
authors.affiliation: Symbiota, Inc, Cambridge, Massachusetts.
authors.forename: Yves
authors.initials: Y
authors.lastname: Millet
authors.affiliation: Department of Genetics, Harvard Medical School and Department of Molecular Biology, Massachusetts General Hospital, Boston, Massachusetts.
authors.forename: Frederick M
authors.initials: FM
authors.lastname: Ausubel
authors.affiliation: Department of Genetics, Harvard Medical School and Department of Molecular Biology, Massachusetts General Hospital, Boston, Massachusetts.
authors.forename: Mark
authors.initials: M
authors.lastname: Borowsky
mesh_terms: D017360:Arabidopsis / Q000235:genetics*; D004252:DNA Mutational Analysis / Q000379:methods; D018744:DNA, Plant / Q000737:chemistry / Q000235:genetics*; D005020:Ethyl Methanesulfonate / Q000737:chemistry*; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods*; D016296:Mutagenesis; D009154:Mutation
chemical_list: D018744:DNA, Plant; D005020:Ethyl Methanesulfonate
keywords: ethyl methanesulfonate (EMS); genetic mapping; next-gen sequencing