title: Identification of an EMS-induced causal mutation in a gene required for boron-mediated root development by low-coverage genome re-sequencing in Arabidopsis.
abstract: Next-generation sequencing (NGS) technologies enable the rapid production of an enormous quantity of sequence data. These powerful new technologies allow the identification of mutations by whole-genome sequencing. However, most reported NGS-based mapping methods, which are based on bulked segregant analysis, are costly and laborious. To address these limitations, we designed a versatile NGS-based mapping method that consists of a combination of low- to medium-coverage multiplex SOLiD (Sequencing by Oligonucleotide Ligation and Detection) and classical genetic rough mapping. Using only low to medium coverage reduces the SOLiD sequencing costs and, since just 10 to 20 mutant F 2 plants are required for rough mapping, the operation is simple enough to handle in a laboratory with limited space and funding. As a proof of principle, we successfully applied this method to identify the CTR1, which is involved in boron-mediated root development, from among a population of high boron requiring Arabidopsis thaliana mutants. Our work demonstrates that this NGS-based mapping method is a moderately priced and versatile method that can readily be applied to other model organisms.
authors.affiliation: Graduate School of Science and Technology; Kumamoto University; Kumamoto, Japan.
authors.forename: Ryo
authors.initials: R
authors.lastname: Tabata
authors.affiliation: Department of Applied Biological Chemistry; Graduate School of Agricultural and Life Sciences; University of Tokyo; Tokyo, Japan.
authors.forename: Takehiro
authors.initials: T
authors.lastname: Kamiya
authors.affiliation: Functional Genomics Facility; National Institute for Basic Biology; Okazaki, Japan.
authors.forename: Shuji
authors.initials: S
authors.lastname: Shigenobu
authors.affiliation: Functional Genomics Facility; National Institute for Basic Biology; Okazaki, Japan.
authors.forename: Katsushi
authors.initials: K
authors.lastname: Yamaguchi
authors.affiliation: Department of Biology and IGSP Center for Systems Biology; Duke University; Durham, NC USA.
authors.forename: Masashi
authors.initials: M
authors.lastname: Yamada
authors.affiliation: Division of Evolutionary Biology; National Institute for Basic Biology; Okazaki, Japan; School of Life Science; The Graduate University for Advanced Studies; Okazaki, Japan; ERATO; Japan Science and Technology Agency; Okazaki, Japan.
authors.forename: Mitsuyasu
authors.initials: M
authors.lastname: Hasebe
authors.affiliation: Department of Applied Biological Chemistry; Graduate School of Agricultural and Life Sciences; University of Tokyo; Tokyo, Japan.
authors.forename: Toru
authors.initials: T
authors.lastname: Fujiwara
authors.affiliation: Graduate School of Science and Technology; Kumamoto University; Kumamoto, Japan.
authors.forename: Shinichiro
authors.initials: S
authors.lastname: Sawa
mesh_terms: D017360:Arabidopsis / Q000235:genetics* / Q000378:metabolism; D001895:Boron / Q000378:metabolism*; D002874:Chromosome Mapping; D005020:Ethyl Methanesulfonate; D017343:Genes, Plant; D059014:High-Throughput Nucleotide Sequencing / Q000379:methods; D009154:Mutation; D018517:Plant Roots / Q000254:growth & development / Q000378:metabolism*; D011494:Protein Kinases / Q000235:genetics* / Q000378:metabolism; D017422:Sequence Analysis, DNA / Q000379:methods*
chemical_list: D005020:Ethyl Methanesulfonate; D011494:Protein Kinases; C477753:CTR1 protein, Arabidopsis; D001895:Boron
keywords: Arabidopsis thaliana; boron; low- to medium-coverage sequencing; mutant mapping; next-generation sequencing