title: Optimizing TILLING populations for reverse genetics in Medicago truncatula.
abstract: Medicago truncatula has been widely adopted as a model plant for crop legume species of the Vicieae. Despite the availability of transformation and regeneration protocols, there are currently limited tools available in this species for the systematic investigation of gene function. Within the framework of the European Grain Legumes Integrated Project (http://www.eugrainlegumes.org), chemical mutagenesis was applied to M. truncatula to create two mutant populations that were used to establish a TILLING (targeting induced local lesions in genomes) platform and a phenotypic database, allowing both reverse and forward genetics screens. Both populations had the same M2 line number, but differed in their M1 population size: population 1 was derived from a small M1 population (one-tenth the size of the M2 generation), whereas population 2 was generated by single seed descent and therefore has M1 and M2 generations of equal size. Fifty-six targets were screened, 10 on both populations, and 546 point mutations were identified. Population 2 had a mutation frequency of 1/485 kb, twice that of population 1. The strategy used to generate population 2 is more efficient than that used to generate population 1, with regard to mutagenesis density and mutation recovery. However, the design of population 1 allowed us to estimate the genetically effective cell number to be three in M. truncatula. Phenotyping data to help forward screenings are publicly available, as well as a web tool for ordering seeds at http://www.inra.fr/legumbase.
authors.affiliation: Unité Mixte de Recherche en Génétique et Ecophysiologie des Légumineuses à Graines, Institut National de la Recherche Agronomique, Dijon, France.
authors.forename: Christine
authors.initials: C
authors.lastname: Le Signor
authors.forename: Vincent
authors.initials: V
authors.lastname: Savois
authors.forename: Grégoire
authors.initials: G
authors.lastname: Aubert
authors.forename: Jérôme
authors.initials: J
authors.lastname: Verdier
authors.forename: Marie
authors.initials: M
authors.lastname: Nicolas
authors.forename: Gaelle
authors.initials: G
authors.lastname: Pagny
authors.forename: Françoise
authors.initials: F
authors.lastname: Moussy
authors.forename: Myriam
authors.initials: M
authors.lastname: Sanchez
authors.forename: Dave
authors.initials: D
authors.lastname: Baker
authors.forename: Jonathan
authors.initials: J
authors.lastname: Clarke
authors.forename: Richard
authors.initials: R
authors.lastname: Thompson
mesh_terms: D004252:DNA Mutational Analysis / Q000379:methods*; D018744:DNA, Plant / Q000235:genetics; D030541:Databases, Genetic; D005020:Ethyl Methanesulfonate; D018506:Gene Expression Regulation, Plant; D017343:Genes, Plant; D018745:Genome, Plant; D005838:Genotype; D046913:Medicago truncatula / Q000235:genetics*; D016296:Mutagenesis; D010641:Phenotype
chemical_list: D018744:DNA, Plant; D005020:Ethyl Methanesulfonate