以烟草为生物反应器表达真菌免疫调节蛋白的初探
发布时间：2018-08-04 14:58
【摘要】： 从1989年至今,科学家们从药用真菌中获得了一系列小分子蛋白质家族——真菌免疫调节蛋白(FIP),证实它可通过激活免疫系统中的细胞间相互作用和各种细胞因子的分泌达到抗肿瘤与免疫调节的功能,从而可对抗癌症及相关免疫性疾病。近年来,科学家们努力探索和研究通过生物反应器来研发与生产大量的有医用价值的药物蛋白。利用植物作为药用蛋白的生物反应器,相对于动物、微生物反应系统及其他生产方式有很多优势,可大量生产可供利用的蛋白质。 本实验以烟草这种模式物种为生物反应器,将从灵芝和金针菇中克隆编码FIP的基因构建到植物表达载体pBI121含有rd29A启动子上,并将该载体转化到农杆菌LBA4404中,通过叶盘转化法对77个外植体进行遗传操作,获得了263个转基因材料,筛选出长势良好的转化植株53株,移栽21株。 通过分子生物学和生理学分析方法对21株转基因植株进行检测和鉴定,证明了外源基因已经整合到烟草基因组中。转基因植株移栽后,根据盐诱导rd29A启动子的特点,通过盐诱导法诱导外源FIP基因的表达,绿色荧光蛋白GFP检测和SDS-PAGE电泳分析显示了FIP基因得到表达,并在盐诱导时表达增强。 本实验结果证实了真菌免疫调节蛋白可以在烟草获得表达,为以烟草作为生物反应器生产真菌免疫蛋白FIP提供了材料及技术基础。
[Abstract]:From 1989 to the present, Scientists have obtained a series of small protein families from medicinal fungi, the fungal immunomodulatory protein (FIP), which has shown that it can resist swelling by activating intercellular interactions in the immune system and the secretion of various cytokines. Tumor and immunomodulation function, This can fight cancer and related immune diseases. In recent years, scientists have made great efforts to develop and produce a large number of medically valuable pharmaceutical proteins through bioreactors. The bioreactor which uses plant as medicinal protein has many advantages over animals, microbial reaction system and other production methods, and can produce the available protein in large quantities. In this experiment, tobacco, a model species, was used as a bioreactor. The gene encoding FIP was cloned from Ganoderma lucidum and Flammulina velutipes to plant expression vector pBI121 containing rd29A promoter, and the vector was transformed into Agrobacterium tumefaciens LBA4404. Through genetic manipulation of 77 explants by leaf disc transformation, 263 transgenic materials were obtained, 53 plants with good growth and 21 plants were transplanted. 21 transgenic plants were detected and identified by molecular biology and physiological analysis, which proved that the exogenous genes had been integrated into the tobacco genome. According to the characteristics of salt-induced rd29A promoter, the expression of exogenous FIP gene was induced by salt-induced method. The expression of FIP gene was detected by green fluorescent protein GFP detection and SDS-PAGE electrophoresis, and the expression of FIP gene was enhanced during salt induction. The results showed that fungal immunomodulin could be expressed in tobacco, which provided a material and technical basis for the production of fungal immune protein FIP by using tobacco as a bioreactor.
【学位授予单位】：东北师范大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R392.1
本文编号：2164237
[Abstract]:From 1989 to the present, Scientists have obtained a series of small protein families from medicinal fungi, the fungal immunomodulatory protein (FIP), which has shown that it can resist swelling by activating intercellular interactions in the immune system and the secretion of various cytokines. Tumor and immunomodulation function, This can fight cancer and related immune diseases. In recent years, scientists have made great efforts to develop and produce a large number of medically valuable pharmaceutical proteins through bioreactors. The bioreactor which uses plant as medicinal protein has many advantages over animals, microbial reaction system and other production methods, and can produce the available protein in large quantities. In this experiment, tobacco, a model species, was used as a bioreactor. The gene encoding FIP was cloned from Ganoderma lucidum and Flammulina velutipes to plant expression vector pBI121 containing rd29A promoter, and the vector was transformed into Agrobacterium tumefaciens LBA4404. Through genetic manipulation of 77 explants by leaf disc transformation, 263 transgenic materials were obtained, 53 plants with good growth and 21 plants were transplanted. 21 transgenic plants were detected and identified by molecular biology and physiological analysis, which proved that the exogenous genes had been integrated into the tobacco genome. According to the characteristics of salt-induced rd29A promoter, the expression of exogenous FIP gene was induced by salt-induced method. The expression of FIP gene was detected by green fluorescent protein GFP detection and SDS-PAGE electrophoresis, and the expression of FIP gene was enhanced during salt induction. The results showed that fungal immunomodulin could be expressed in tobacco, which provided a material and technical basis for the production of fungal immune protein FIP by using tobacco as a bioreactor.
【学位授予单位】：东北师范大学
【学位级别】：硕士
【学位授予年份】：2008
【分类号】：R392.1
【引证文献】
相关硕士学位论文 前1条
1 王雪飞;基因重组真菌免疫调节蛋白的筛选与功能检测[D];上海交通大学;2012年
，本文编号：2164237
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