转人β防御素-3基因牛奶食用安全性评价

发布时间：2019-01-03 19:38

【摘要】：随着现代分子生物学、动物胚胎学和配子生物工程技术的发展，多种转基因家畜已经研制成功，在生物基础研究、医学、农业、环境保护等领域已显示了广阔的应用前景。与此同时转基因食品的安全与否也成为了社会各界关注的焦点问题。因此转基因食品安全与否，对于转基因技术的发展、转基因食品的推广以及解决全球的粮食短缺问题都具有重要意义。本研究结合FDA/WHO发布的实质等同原则、植物转基因安全研究的先例和农业部关于转基因食品安全的国家标准及食品安全标准，从转β防御素3基因牛奶营养成分分析，外源蛋白生物信息学分析和转基因食品饲喂小鼠食用安全性实验三方面入手，评价转基因牛奶的食用安全性。本研究的实施为转基因动物产品的后期应用提供了一定的实验和理论依据，为建立转基因动物及其产品安全性检测体系进行了初步的探索。 1.牛奶营养成分分析本实验以转人β防御素3基因牛奶为实验样品，以普通牛奶作为对照，对比了两种牛奶的基本营养成分（包括脂肪、总蛋白含量、乳糖含量、PH、非脂乳固体）、氨基酸、主要矿质元素（包括Ca、Mg、P、K、Na、Fe、Zn、Se）、维生素（包括维生素A、维生系E、维生素C、维生素B1、维生素B2）的含量。实验结果表明，，转人β防御素3基因牛奶在营养成分方面与普通牛奶比较，其差异没有统计学意义。 2.生物信息学分析使用ExPASy网站中的Pepetide Mass、ProtParam tool和Portscale对人β防御素3进行蛋白的结构和理化分析。使用SDAP和Farrp数据库对人β防御素3与致敏性蛋白的同源序列对比；使用PDB和PHYRE数据库进行三维结构管比对，人β防御素3的致敏性。实验结果：1)人β防御素3不是来自于公认的八类主要致敏原；2）理化性质预测显示人β防御素3是一种不稳定的蛋白质；3）生物信息学分析显示人β防御素3与数据库中的致敏性蛋白没有同源性序列，提示β防御素3不具有致敏性。 3.转人β防御素基因牛奶小鼠经口30天饲喂实验本实验将小鼠分为三组，分别为灌胃饲喂转人β防御素3基因牛奶组、普通牛奶组、空白对照组。各实验组小鼠饲喂30天后，腹腔麻醉，经心脏采血分别用于血液学常规检查和生化指标检测。采血后，取小鼠脑、心脏、肺脏、肝脏、脾脏、胃、大肠、小肠、肾脏等主要脏器组织制作切片后进行HE染色。实验结果显示，各实验组血液学指标、血清生化指标无显著差异，大体解剖和病理学检查未见异常。
[Abstract]:With the development of modern molecular biology, animal embryology and gametes bioengineering, many kinds of transgenic livestock have been successfully developed, and have shown a broad application prospect in biological basic research, medicine, agriculture, environmental protection and other fields. At the same time, the safety of GM food has become the focus of attention. Therefore, the safety or not of genetically modified food is of great significance to the development of transgenic technology, the promotion of transgenic food and the solution of global food shortage. Based on the principle of substantial equivalence published by FDA/WHO, the precedent of plant transgenic safety research and the national standards and food safety standards of the Ministry of Agriculture on the safety of genetically modified foods, the nutritional composition of milk with 尾 -defensin 3 gene was analyzed. The food safety of transgenic milk was evaluated by bioinformatics analysis of exogenous protein and experiment of food safety of transgenic food feeding mice. The implementation of this study provides a certain experimental and theoretical basis for the later application of transgenic animal products, and provides a preliminary exploration for the establishment of a safety detection system for transgenic animals and their products. 1. Nutritional composition Analysis of Milk in this study, the basic nutritional components (including fat, total protein content, lactose content, PH,) of two kinds of milk were compared using human 尾 -defensin 3 gene milk as experimental sample and ordinary milk as control. Non-fat milk solid), amino acids, main mineral elements (including Ca,Mg,P,K,Na,Fe,Zn,Se), vitamins (including vitamin A, life line E, vitamin C, vitamin B1, vitamin B2). The results showed that there was no significant difference in nutritional composition of human 尾 -defensin 3 gene milk compared with that of common milk. 2. Bioinformatics analysis of human 尾-defensin 3 protein using Pepetide Mass,ProtParam tool and Portscale in ExPASy website. The homologous sequences of human 尾 -defensin 3 and sensitized protein were compared by using SDAP and Farrp databases, and the sensitivities of human 尾 -defensin 3 were obtained by using PDB and PHYRE databases for three-dimensional structural tube alignment. The results were as follows: 1) Human 尾 -defensin 3 was not derived from eight major allergens, 2) the physical and chemical properties predicted that human 尾 -defensin 3 was an unstable protein. 3) Bioinformatics analysis showed that human 尾 -defensin 3 had no homologous sequence with the sensitized protein in the database, suggesting that 尾 -defensin 3 was not sensitized. 3. The mice were divided into three groups: human 尾 -defensin 3 gene milk group, normal milk group and blank control group. After 30 days of feeding, the mice in each experimental group were anesthetized intraperitoneally, and blood was collected from the heart for routine hematological examination and biochemical index detection. After blood collection, the brain, heart, lung, liver, spleen, stomach, large intestine, small intestine, kidney and other main organs of the mice were taken for HE staining. The results showed that there were no significant differences in hematological and serum biochemical indexes among the experimental groups, and there were no abnormalities in gross anatomy and pathology.
【学位授予单位】：西北农林科技大学
【学位级别】：硕士
【学位授予年份】：2014
【分类号】：TS252.1

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