A high-density collection of EMS-induced mutations for TILLING in Landsberg erecta genetic background of Arabidopsis View Full Text


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Article Info

DATE

2009-12

AUTHORS

Beatriz Martín, Mercedes Ramiro, José M Martínez-Zapater, Carlos Alonso-Blanco

ABSTRACT

BACKGROUND: Arabidopsis thaliana is the main model species for plant molecular genetics studies and world-wide efforts are devoted to identify the function of all its genes. To this end, reverse genetics by TILLING (Targeting Induced Local Lesions IN Genomes) in a permanent collection of chemically induced mutants is providing a unique resource in Columbia genetic background. In this work, we aim to extend TILLING resources available in A. thaliana by developing a new population of ethyl methanesulphonate (EMS) induced mutants in the second commonest reference strain. In addition, we pursue to saturate the number of EMS induced mutations that can be tolerated by viable and fertile plants. RESULTS: By mutagenizing with different EMS concentrations we have developed a permanent collection of 3712 M2/M3 independent mutant lines in the reference strain Landsberg erecta (Ler) of A. thaliana. This population has been named as the Arabidopsis TILLer collection. The frequency of mutations per line was maximized by using M1 plants with low but sufficient seed fertility. Application of TILLING to search for mutants in 14 genes identified 21 to 46 mutations per gene, which correspond to a total of 450 mutations. Missense mutations were found for all genes while truncations were selected for all except one. We estimated that, on average, these lines carry one mutation every 89 kb, Ler population providing a total of more than five million induced mutations. It is estimated that TILLer collection shows a two to three fold higher EMS mutation density per individual than previously reported A. thaliana population. CONCLUSIONS: Analysis of TILLer collection demonstrates its usefulness for large scale TILLING reverse genetics in another reference genetic background of A. thaliana. Comparisons with TILLING populations in other organisms indicate that this new A. thaliana collection carries the highest chemically induced mutation density per individual known in diploid species. More... »

PAGES

147

References to SciGraph publications

  • 1986-06. Genetics of actin-related sequences in tomato in THEORETICAL AND APPLIED GENETICS
  • 2007-12. Discovery of chemically induced mutations in rice by TILLING in BMC PLANT BIOLOGY
  • 2003-09. An Arabidopsis thaliana T-DNA mutagenized population (GABI-Kat) for flanking sequence tag-based reverse genetics in PLANT MOLECULAR BIOLOGY
  • 2006-12. A protocol for TILLING and Ecotilling in plants and animals in NATURE PROTOCOLS
  • 2005-09. Reverse Genetic Approaches for Functional Genomics of Rice in PLANT MOLECULAR BIOLOGY
  • 2006-12. TILLING is an effective reverse genetics technique for Caenorhabditis elegans in BMC GENOMICS
  • 2009-03. Forward and reverse genetics of rapid-cycling Brassica oleracea in THEORETICAL AND APPLIED GENETICS
  • 2008-12. Applying genotyping (TILLING) and phenotyping analyses to elucidate gene function in a chemically induced sorghum mutant population in BMC PLANT BIOLOGY
  • 2008-12. TILLING to detect induced mutations in soybean in BMC PLANT BIOLOGY
  • 2005-01. A reverse genetic, nontransgenic approach to wheat crop improvement by TILLING in NATURE BIOTECHNOLOGY
  • 2004-12. Discovery of induced point mutations in maize genes by TILLING in BMC PLANT BIOLOGY
  • 2008-03. MNU-induced mutant pools and high performance TILLING enable finding of any gene mutation in rice in MOLECULAR GENETICS AND GENOMICS
  • 2000-12. Analysis of the genome sequence of the flowering plant Arabidopsis thaliana in NATURE
  • 2008-12. An Italian functional genomic resource for Medicago truncatula in BMC RESEARCH NOTES
  • Identifiers

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    http://scigraph.springernature.com/pub.10.1186/1471-2229-9-147

    DOI

    http://dx.doi.org/10.1186/1471-2229-9-147

    DIMENSIONS

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    PUBMED

    https://www.ncbi.nlm.nih.gov/pubmed/20003424


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    43 schema:description BACKGROUND: Arabidopsis thaliana is the main model species for plant molecular genetics studies and world-wide efforts are devoted to identify the function of all its genes. To this end, reverse genetics by TILLING (Targeting Induced Local Lesions IN Genomes) in a permanent collection of chemically induced mutants is providing a unique resource in Columbia genetic background. In this work, we aim to extend TILLING resources available in A. thaliana by developing a new population of ethyl methanesulphonate (EMS) induced mutants in the second commonest reference strain. In addition, we pursue to saturate the number of EMS induced mutations that can be tolerated by viable and fertile plants. RESULTS: By mutagenizing with different EMS concentrations we have developed a permanent collection of 3712 M2/M3 independent mutant lines in the reference strain Landsberg erecta (Ler) of A. thaliana. This population has been named as the Arabidopsis TILLer collection. The frequency of mutations per line was maximized by using M1 plants with low but sufficient seed fertility. Application of TILLING to search for mutants in 14 genes identified 21 to 46 mutations per gene, which correspond to a total of 450 mutations. Missense mutations were found for all genes while truncations were selected for all except one. We estimated that, on average, these lines carry one mutation every 89 kb, Ler population providing a total of more than five million induced mutations. It is estimated that TILLer collection shows a two to three fold higher EMS mutation density per individual than previously reported A. thaliana population. CONCLUSIONS: Analysis of TILLer collection demonstrates its usefulness for large scale TILLING reverse genetics in another reference genetic background of A. thaliana. Comparisons with TILLING populations in other organisms indicate that this new A. thaliana collection carries the highest chemically induced mutation density per individual known in diploid species.
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