薏苡仁油含量与相关候选基因的关联分析

Coix seed is a valuable traditional chinese medicine and health food, in which Coix seed oil is one of the main active ingredients with anti-tumor and hypolycemic effect and has been developed for injection that was widely used in the treatment of various clinical cancer，therefore, it is significance that studying improving the oil content or altering the oil composition of Coix seed. In this study, the content of Coix seed oil and triglyceride in 298 Coix lacryma-jobi L. from Guizhou, Yunnan, Sichuan and Guangxi were tested, the variation of Coix seed oil and triglyceride in the population was analyzed, the selected germplasm with high Coix seed oil content could be used as parent material in breeding program for breeding high oil content variety. Three genomic sequences of genes encoding Acyl-CoA diacylglycerol acyltransferase (DGAT), Stearoyl-ACP desaturase (SAD) and ER oleate desaturase (FAD2) were screened among the 298 Coix lacryma-jobi L. and analyzed for nucleotide polymorphisms and linkage disequilibrium, respectively. Combined with population structure and kinship, association analysis were then carried out using single nucleotide polymorphism and Coix seed oil content dat. Loci associated with Coix seed oil obtained in this study lays the foundation for developing functional makers associated with high Coix seed oil and will be useful for the molecular breeding of high Coix seed oil vavieties in the future.

薏苡仁是药食兼用的珍贵中药和保健食品，薏苡仁油是主要活性成分之一，具有抗肿瘤和降血糖功效，已被研发为注射液广泛用于临床各种癌症的治疗，开展提高薏苡仁油含量或改变成分组成的研究具有重要意义。本研究对来源于我国贵州、云南、四川和广西4省的298份薏苡资源进行薏苡仁油及甘油三酯含量的检测，分析薏苡仁油及甘油三酯含量在群体中的变异幅度，获得的高薏苡仁油含量种质可作为高油薏苡育种的亲本资源。分析薏苡酰基辅酶A二酰甘油酰基转移酶(DGAT)、硬脂酸脱饱和酶(SAD)和内质网油酸脱饱和酶(FAD2)3个基因的基因组序列在298份薏苡资源中的核苷酸多态性和连锁不平衡，结合群体结构和亲缘关系，薏苡仁油及甘油三酯含量数据，进行关联分析，获得与薏苡仁油性状相关的遗传位点，为开发高薏苡仁油相关功能标记奠定基础，有利于后期开展高油薏苡分子育种。

薏苡仁是我国古老的珍贵药材和保健食品，薏苡仁油是其主要的活性成分。本项目对我国西南地区广泛收集的薏苡仁脂肪酸和甘油三酯的含量进行了检测，发现10种游离脂肪酸在所有样本中均未检测到，4种游离脂肪酸仅在少数几个样本中检测到，有21种脂肪酸可以在绝大数样本中检测到，并表现出广泛变异，含量最大的4种脂肪酸为油酸，平均含量为2.34g/100g，亚油酸平均为1.79g/100g，棕榈酸平均为0.71g/100g，硬脂酸平均为0.13g/100g，含量最小的脂肪酸为花生四烯酸，平均含量为0.0013g/100g。另外，除了亚油酸和花生四烯酸外，薏苡籽粒油脂中还含有人体不能合成的脂肪酸，如α-亚麻酸，γ-亚麻酸，仅在3个样本中检测到了二十碳五烯酸（EPA），在1个样本中检测到了二十二碳六烯酸（DHA）。5种甘油三酯含量检测结果表明，三亚油酸甘油酯为3228.87mg/kg，1,2-亚油酸-3-油酸甘油酯含量为12637.68mg/kg，1,2-油酸-3-亚油酸甘油酯含量为11909.60mg/kg，1-棕榈酸-2-油酸-3-亚油酸甘油酯含量为42942.01mg/kg，三油酸甘油酯含量为13501.93mg/kg，因此，1-棕榈酸-2-油酸-3-亚油酸甘油酯含量最高，三亚油酸甘油酯含量最低。克隆了薏苡油脂合成关键基因FAD2和SAD的基因组序列，长度分别为1189bp和4288bp，ClFAD2和ClSAD基因启动子序列上均含有抗逆性相关顺式作用元件和植物激素响应元件。两个基因在我国西南地区薏苡种质中序列多样性检测结果表明，FAD2存在3个SNPs位点，无Indel标记，SAD基因可找到15个SNPs位点和1个长259bp的Indel位点。开发了薏苡关联群体Indel标记124754个和SNP标记724728个，分析表明，薏苡种质亲缘关系在0-0.15之间，可分为8个亚群。利用关联分析研究策略，获得了薏苡仁脂肪酸组分和5种甘油三酯含量相关的显著性遗传标记。本项目研究结果不仅筛选鉴定了一批富含油脂或功能性成分的薏苡种质，为高油或功能性薏苡品种的选育提供了种质基础，同时，薏苡籽粒油脂含量遗传标记的获得，有助于利用分子标记辅助选择并结合传统育种方法将高油等位基因导入普通高产薏苡材料，加速高薏苡仁油品种的选育进程。