title: A novel viable allele of Arabidopsis CULLIN1 identified in a screen for superroot2 suppressors by next generation sequencing-assisted mapping.
abstract: Map-based cloning (MBC) is the conventional approach for linking phenotypes to genotypes, and has been successfully used to identify causal mutations in diverse organisms. Next-generation sequencing (NGS) technologies offer unprecedented possibilities to sequence the entire genomes of organisms, thereby in principle enabling direct identification of causal mutations without mapping. However, although mapping-by-sequencing has proven to be a cost effective alternative to classical MBC in particular situations, methods based solely on NGS still have limitations and need to be refined. Aiming to identify the causal mutations in suppressors of Arabidopsis thaliana superroot2 phenotype, generated by ethyl methane sulfonate (EMS) treatment, we combined NGS and classical mapping, to rapidly identify the point mutations and restrict the number of testable candidates by defining the chromosomal intervals containing the causal mutations, respectively. The NGS-assisted mapping approach we describe here facilitates unbiased identification of virtually any causal EMS-generated mutation by overlapping the identification (deep sequencing) and validation (mapping) steps. To exemplify the useful marriage of the two approaches we discuss the strategy used to identify a new viable recessive allele of the Arabidopsis CULLIN1 gene in the non-reference Wassilewskija (Ws-4) accession.
authors: Department of Plant Physiology, Umeå Plant Science Centre, Umeå University, Umeå, Sweden.
authors: Daniel I
authors: DI
authors: Pacurar
authors: Department of Plant Physiology, Umeå Plant Science Centre, Umeå University, Umeå, Sweden; Faculty of Horticulture, University of Agricultural Sciences and Veterinary Medicine, Cluj Napoca, Romania; Present address: SweTree Technologies AB, Umeå, Sweden.
authors: Monica L
authors: ML
authors: Pacurar
authors: Faculty of Horticulture, University of Agricultural Sciences and Veterinary Medicine, Cluj Napoca, Romania.
authors: Andrea M
authors: AM
authors: Pacurar
authors: Molecular biology platform (CRRBM), Université de Picardie Jules Verne, Amiens, France.
authors: Laurent
authors: L
authors: Gutierrez
authors: Department of Plant Physiology, Umeå Plant Science Centre, Umeå University, Umeå, Sweden; Institut Jean-Pierre Bourgin, French National Institute for Agricultural Research (UMR1318 INRA-AgroParisTech), Versailles, France.
authors: Catherine
authors: C
authors: Bellini
mesh_terms: D000483:Alleles; D017360:Arabidopsis / Q000235:genetics*; D002874:Chromosome Mapping; D032461:Chromosomes, Plant / Q000235:genetics; D044842:Cullin Proteins / Q000235:genetics* / Q000378:metabolism; D005020:Ethyl Methanesulfonate; D005808:Genes, Recessive; D016153:Genes, Suppressor; D005820:Genetic Testing; D059014:High-Throughput Nucleotide Sequencing; D058977:Molecular Sequence Annotation; D009154:Mutation / Q000235:genetics; D010641:Phenotype; D036226:Seedlings / Q000235:genetics
chemical_list: C100318:Cullin 1; D044842:Cullin Proteins; D005020:Ethyl Methanesulfonate