Development of low-linolenic acid Brassica oleracea lines through seed mutagenesis and molecular characterization of mutants
Designing the fatty acid composition of Brassica napus L. seed oil for specific applications would extend the value of this crop. A mutation in Fatty Acid Desaturase 3 (FAD3), which encodes the desaturase responsible for catalyzing the formation of α-linolenic acid (ALA; 18:3 cisΔ9,12,15), in a diploid Brassica species would potentially result in useful germplasm for creating an amphidiploid displaying low ALA content in the seed oil. For this, seeds of B. oleracea (CC), one of the progenitor species of B. napus, were treated with ethyl-methane-sulfonate to induce mutations in genes encoding enzymes involved in fatty acid biosynthesis. Seeds from 1,430 M2 plants were analyzed, from which M3 seed families with 5.7–6.9 % ALA were obtained. Progeny testing and selection for low ALA content were carried out in M3–M7 generations, from which mutant lines with <2.0 % ALA were obtained. Molecular analysis revealed that the mutation was due to a single nucleotide substitution from G to A in exon 3 of FAD3, which corresponds to an amino acid residue substitution from glutamic acid to lysine. No obvious differences in the expression of the FAD3 gene were detected between wild type and mutant lines; however, evaluation of the performance of recombinant Δ-15 desaturase from mutant lines in yeast indicated reduced production of ALA. The novelty of this mutation can be inferred from the position of the point mutation in the C-genome FAD3 gene when compared to the position of mutations reported previously by other researchers. This B. oleracea mutant line has the potential to be used for the development of low-ALA B. napus and B. carinata oilseed crops.
KeywordsMutant Line Napus Line Tyloxapol Produce Fatty Acid Methyl Ester Sixth Intron
HR and RJW are grateful for the support provided by AVAC Ltd, the Canada Foundation for Innovation and the Research Capacity Program of Alberta Enterprise and Advanced Education. RJW is also grateful for the support provided by the Alberta Innovates Bio Solutions, Canada Research Chairs Program and the Natural Sciences and Engineering Research Council of Canada. The authors also thank Dr. Mohan Thiagarajah for suggestions on EMS treatments, Dr. Nidhi Sharma for collecting developing siliques, Ms. An Vo for FA analysis of the mutagenized populations, and other laboratory staff for the technical assistance provided.
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