Flores Teresita, Karpova Olga, Su Xiujuan, Zeng Peiyu, Bilyeu Kristin, Sleper David A, Nguyen Henry T, Zhang Zhanyuan J
Plant Transformation Core Facility, Division of Plant Sciences, University of Missouri, 1-31 Agriculture Building, Columbia, MO, 65211-7140, USA.
Transgenic Res. 2008 Oct;17(5):839-50. doi: 10.1007/s11248-008-9167-6. Epub 2008 Feb 7.
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 alpha-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 alpha-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.
RNA干扰（RNAi）最近已被用作一种有效的实验工具，用于包括植物在内的各种生物体的基础生物学研究和应用生物学研究。RNAi利用小RNA，主要是小干扰RNA（siRNA），来介导mRNA的降解，从而调节植物中的基因表达。在此，我们报道了在复杂基因组大豆（Glycine max）中，通过siRNA介导的ω-3脂肪酸去饱和酶（FAD3）基因家族的高效基因沉默。FAD3酶负责多不饱和脂肪酸途径中α-亚麻酸（18:3）的合成。正是这种脂肪酸在很大程度上导致了大豆和其他种子油的不稳定性。因此，这种脂肪酸的显著减少将提高种子油的稳定性，增强种子的农艺价值。一个长度为318个核苷酸的保守核苷酸序列，为这三个基因家族成员所共有，被用作RNA干扰的反向重复序列。RNAi表达盒由种子特异性启动子驱动。我们表明，转基因产生的siRNA导致FAD3沉默，其效果与fad3突变体表型相当，此外，这种沉默在转基因大豆品系中能够稳定遗传。由于相对于大豆中的其他多不饱和脂肪酸，α-亚麻酸的总量较少，这种脂肪酸的显著减少表明siRNA策略在复杂基因组中沉默基因家族方面具有作用和巨大潜力。
