紫花苜蓿FAD基因调控α-亚麻酸生物合成机制研究

α-linolenic acid is essential for human health, and its deficiency in the diet is a major explanation for many diseases. Alfalfa (Medicago sativa subsp. sativa) is the most widely cultivated leguminous forage in the world, but its low content of α-linolenic acid directly affects the α-linolenic acid proportions and nutritional value of milk. The regulatory mechanism of α-linolenic acid biosynthesis in alfalfa is not clear to date. Δ12 fatty acid desaturases (FADs) and Δ15FADs are the key enzymes involved in the biosynthetic pathway of α-linolenic acid. Through high-throughput transcriptome sequencing and RACE methods, the full-length of 2 Δ12FAD and 2 Δ15FAD cDNA sequences were cloned from alfalfa. Based on this, we aim to analyze the expression pattern, subcellular localization and biological function of Δ12FAD and Δ15FAD genes, clarify the key genes and the regulatory mechanism of the α-linolenic acid biosynthesis in alfalfa, and create materials with high content of α-linolenic acid. The results in this research can provide important theoretical foundations for the improvement of alfalfa quality and nutritional value of dairy products.

α-亚麻酸是人体必需脂肪酸，长期摄入不足极易诱发多种疾病。紫花苜蓿是全球种植面积最大的豆科牧草，但其α-亚麻酸含量低，直接影响乳产品α-亚麻酸比例及其营养价值。紫花苜蓿α-亚麻酸生物合成机制仍不清楚。Δ12脂肪酸去饱和酶（FADs）和Δ15FADs是调控α-亚麻酸生物合成途径的关键酶。通过高通量转录组测序和RACE方法，申请人从紫花苜蓿中克隆了2个Δ12FAD和2个Δ15FAD基因的cDNA全长。在此基础上，本研究拟通过系统分析紫花苜蓿Δ12FAD和Δ15FAD基因的表达模式、亚细胞定位和生物学功能，明确调控紫花苜蓿叶片α-亚麻酸生物合成的关键基因，解析紫花苜蓿α-亚麻酸生物合成调控机制，获得高含量α-亚麻酸紫花苜蓿材料。该项目的实施和完成，可为改良紫花苜蓿品质和提高乳产品营养价值提供重要理论基础。

α-亚麻酸是人体必需脂肪酸，对于人体健康及大脑发育具有重要作用。由于人体自身不能合成α-亚麻酸，必须从外界食物中获取。因此，深入解析动植物α-亚麻酸分子代谢途径机理对于缓解人体多不饱和脂肪酸摄取不足，增进人体健康以及实现医食同源都具有重要意义。本项目利用生物信息学方法，在蒺藜苜蓿（Medicago truncatula）和紫花苜蓿（Medicago sativa subsp. sativa）全基因组中共分别鉴定出20个蒺藜苜蓿FADs基因家族成员和33个紫花苜蓿FADs基因家族成员。在蒺藜苜蓿中，MtFAD3.1和MtFAD3.2基因受高温胁迫诱导表达，而MtFAD7.1和MtFAD7.2则响应低温胁迫，表明MtFAD3和MtFAD7基因在响应温度胁迫中发挥功能。紫花苜蓿MsFAD3和MsFAD6基因烟草表皮亚细胞定位结果表明，MtFAD3和MtFAD6分别定位在内质网上和叶绿体上。在紫花苜蓿中过表达MtFAD3和MtFAD6后可显著提高紫花苜蓿叶片中亚麻酸含量，并获得了19株亚麻酸高含量转基因株系。基于以上研究结果，目前已发表学术论文10篇（SCI论文6篇），申请国家发明专利2项，参编英文学术专著1部，培养青年骨干1名，研究生5名，超额完成了预期目标。