基于基因编辑技术创制高亚油酸水稻材料

doi:10.3864/j.issn.0578-1752.2021.14.001

摘要/Abstract

摘要：

【目的】亚油酸是一种人体必需的脂肪酸,能加快体内脂肪代谢与分解、减少胆固醇在血管壁上形成积存、有效预防动脉硬化的发生、提高人体免疫力、促进骨骼发育、提高记忆力、预防脑部功能退化等功能。利用基因编辑技术对控制脂肪酸合成途径的关键酶ω-3脂肪酸去饱和酶基因进行精准编辑,关闭下游产物的合成途径,以便提高水稻上游亚油酸的含量,为创制富集亚油酸水稻材料提供依据。【方法】水稻脂肪酸合成途径中的关键酶ω-3脂肪酸去饱和酶基因（OsFAD3）存在2个拷贝,分别分布在第11和第12染色体,且两者cDNA同源性达97.32%。根据基因编辑原理（编辑的特异性取决于引导RNA（gRNA）的特性）,在2个同源基因序列高度一致的外显子区域,分别设计合成2个gRNA,并分别构建植物基因编辑载体,利用农杆菌介导法转化本地受体材料（富源4号）。通过对基因编辑植株进行靶位点测序鉴定,分析2个位点的编辑效率和基因型。同时,对OsFAD3双突变体进行粒宽、粒长等主要籽粒农艺性状分析。采用气质联用检测法测量OsFAD3双突变体籽粒的37种脂肪酸含量。【结果】获得2个位点的纯合编辑材料,同时也获得了其他不同基因型组合的编辑材料;与对照材料相比,OsFAD3双突变体的粒宽、粒长等主要籽粒农艺性状均没有发生显著变化,但稻谷中亚油酸的相对含量提高了3.36个百分点。【结论】在不改变籽粒主要农艺性状的前提下,实现了利用基因编辑技术通过转化一个载体同时精准敲除2个ω-3脂肪酸去饱和酶基因,提高了籽粒中亚油酸的相对含量。

关键词: 水稻, 基因编辑, 高亚油酸, 米糠油

Abstract:

【Objective】 Linoleic acid is one of essential fatty acid for human, which can accelerate the metabolism and decomposition of fat in the body, reduce the accumulation of cholesterol on the wall of blood vessels, effectively prevent the occurrence of arteriosclerosis, improve human immunity, promote bone development, improve memory and prevent brain function degradation. Rice bran is a by-product of rice processing, and is an edible oil with high nutritional value. In this paper, we precisely edited the gene of ω-3 fatty acid desaturase, which is the key enzyme controlling fatty acid synthesis pathway. It can enrich the upstream linoleic acid, while closing the synthesis pathway of downstream products. 【Method】There are two copies of the key enzyme ω-3 fatty acid desaturase gene ( OsFAD3) in the fatty acid synthesis pathway in rice, which are located on chromosome 11 and chromosome 12, and their cDNA homology is 97.32%. According to the basic principles of gene editing, the specificity of edit depends on the characteristics of the guide RNA (gRNA). In this study, two gRNAs were designed and synthesized in the exon region of two homologous genes, and were constructed into editing vectors respectively. The local receptor material (Fuyuan 4) was successfully transformed by Agrobacterium mediated method. The editing efficiency and genotype of the two sites were analyzed. At the same time, the main agronomic characters of OsFAD3 double mutant were measured, such as grain width and grain length. The contents of 37 fatty acids were figured by GC-MS in the grains of OsFAD3double mutants. 【Result】The results showed two homozygous editing materials were obtained, and other editing materials with different genotypes were obtained. Compared with the control materials, the main agronomic traits such as grain width and grain length of OsFAD3 double mutant materials had no significant changes, however, the relative content of linoleic acid in rice increased by 3.36%. 【Conclusion】One gRNA vector knock out two ω-3 fatty acid desaturase genes at the same time, which improves the relative content of linoleic acid in grains without changing the main agronomic traits of seeds.

Key words: rice, gene edit, high linoleic, rice bran

陈晓军,吴凯伸,惠建,白海波,马斯霜,李靖宇. 基于基因编辑技术创制高亚油酸水稻材料[J]. 中国农业科学, 2021, 54(14): 2931-2940.

CHEN XiaoJun,WU KaiShen,HUI Jian,BAI HaiBo,MA SiShuang,LI JingYu. Production of High Linoleic Acid Rice by Genome Editing[J]. Scientia Agricultura Sinica, 2021, 54(14): 2931-2940.

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https://www.chinaagrisci.com/CN/Y2021/V54/I14/2931

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