Silencing of GmFAD3 gene by siRNA leads to low α-linolenic acids (18:3) of fad3-mutant phenotype in soybean [Glycine max (Merr.)]
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RNA interference (RNAi) has been recently employed as an effective experimental tool for both basic and applied biological studies in various organisms including plants. RNAi deploys small RNAs, mainly small interfering RNAs (siRNAs), to mediate the degradation of mRNA for regulating gene expression in plants. Here we report an efficient siRNA-mediated gene silencing of the omega-3 fatty acid desaturase (FAD3) gene family in a complex genome, the soybean (Glycine max). The FAD3 enzyme is responsible for the synthesis of α-linolenic acids (18:3) in the polyunsaturated fatty acid pathway. It is this fatty acid that contributes mostly to the instability of soybean and other seed oils. Therefore, a significant reduction of this fatty acid will increase the stability of the seed oil, enhancing the seed agronomical value. A conserved nucleotide sequence, 318-nt in length, common to the three gene family members was used as an inverted repeat for RNA interference. The RNAi expression cassette was driven by a seed-specific promoter. We show that the transgene-produced siRNA caused silencing of FAD3 that was comparable to the fad3 mutant phenotype and, furthermore, that such a silencing is stably inherited in engineered soybean lines. Since the pool size of the α-linolenic acids is small relative to the other polyunsaturated fatty acids in soybean, the significant reduction of this fatty acid suggests a role and great potential for the siRNA strategy in silencing gene families in a complex genome.
KeywordsRNAi siRNA Inverted repeat Gene silencing GmFAD3
We thank Christine Cole (University of Missouri-Columbia) for FA analysis of soybean seeds; James Birchler and Seth D. Findley (University of Missouri-Columbia) and Amitava Mitra (University of Nebraska-Lincoln) for critical review of the manuscript; Richard Jorgensen and Vicki Chandler (University of Arizona) for vector pMCG161; James Carrington (Oregon State University) for vector pRTL2; Edgar Cahoon (Donald Danforth Plant Science Center) for the soybean glycinin seed-specific promoter, and Amitava Mitra (University of Nebraska-Lincoln) for helpful discussions. This work was supported by University of Missouri-Columbia Life Science Mission Enhancement program and, in part, by the Illinois-Missouri Biotechnology Alliance (special USDA grant) and Missouri Soybean Merchandizing Council.
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