反义RNA介导GmFAD2-1B基因沉默增强大豆种子中油酸的高效累积

摘要

油酸含量是评价大豆油食用品质和稳定性的重要指标.本研究采用农杆菌介导转化法, 将反义GmFAD2-1B基因导入栽培大豆品种,获得油酸含量显著提高的转基因大豆新品系.Southern杂交检测表明,外源GmFAD2-1B基因片段已导入大豆基因组,其插入拷贝数为1~5个.qRT-PCR检测表明,外源GmFAD2-1B主要在大豆种子中表达,并导致种子中内源GmFAD2-1 mRNA表达水平显著降低,而根、茎、叶、花组织中内源GmFAD2-1 mRNA表达水平无显著变化.脂肪酸组分分析表明, 12份转基因大豆种子油酸含量为27.38%~80.42%, 其中, L40和L72油酸含量分别为68.91%~80.42%和65.98%~80.22%, 较对照品种 Williams 82 (17.8%~22.0%)提高2.65倍以上, 亚油酸降低至4.84%~14.55%, 饱和脂肪酸降低至10.34%~11.16%, 但总脂肪和总蛋白含量与对照品种相比没有显著变化.农艺性状分析表明, 转基因大豆在熟期、株高、叶形、花色、结荚高度、百粒重等方面与对照品种也没有显著差异.%The content of oleic acid is usually taken as a crucial indicator of cooking quality and stability for soybean oil. Breed-ing soybean cultivars with high oleic acid has always been considered as one of the principal aims for soybean improvement. In this study, the anti-sense GmFAD2-1B gene from soybean was transformed into the cultivated soybean by Agrobacte-rium-mediated method and several soybean transgenic lines with dramatically increased oleic acid content were obtained.South-ern blot analysis confirmed integration of the anti-sense GmFAD2-1B fragment into the soybean genome with one to five copies of T-DNA insertions in the selected independent transgenic plants. The results of qRT-PCR showed that the foreign anti-sense GmFAD2-1B was mainly expressed in the transgenic seeds and simultaneously resulted in a significant decrease of the endoge-nous GmFAD2-1 mRNA,however,there were no significant differences observed for GmFAD2-1 mRNA contents in the root,leaf, stem and flower tissues of the transgenic plants compared to the non-transgenic ones. Analysis of the fatty acid compositions in-dicated that the content of oleic acid increased to 27.38%-80.42% in twelve transgenic lines. Specifically, in transgenic lines L40 and L72, oleic acid content was raised to 68.91%-80.42% and 65.98%-80.22% respectively, with over 2.65-fold increase com-pared with the wild type Williams 82 (containing 17.8%-22.0% of oleic oil). Concomitantly, linoleic acid and saturated fatty acid were reduced to 4.84%-14.55% and 10.34%-11.16% respectively in the seeds of transgenic lines L40 and L72. However, there was no significant difference in total oil and protein contents, as well as agronomic traits such as maturity period, plant height, leaf shape, flower color, pod height and weight per 100 seeds between the transgenic lines and non-transformed controls.