【摘要】： 從1989年至今,科學家們從藥用真菌中獲得了一系列小分子蛋白質(zhì)家族——真菌免疫調(diào)節(jié)蛋白(FIP),證實它可通過激活免疫系統(tǒng)中的細胞間相互作用和各種細胞因子的分泌達到抗腫瘤與免疫調(diào)節(jié)的功能,從而可對抗癌癥及相關免疫性疾病。近年來,科學家們努力探索和研究通過生物反應器來研發(fā)與生產(chǎn)大量的有醫(yī)用價值的藥物蛋白。利用植物作為藥用蛋白的生物反應器,相對于動物、微生物反應系統(tǒng)及其他生產(chǎn)方式有很多優(yōu)勢,可大量生產(chǎn)可供利用的蛋白質(zhì)。 本實驗以煙草這種模式物種為生物反應器,將從靈芝和金針菇中克隆編碼FIP的基因構(gòu)建到植物表達載體pBI121含有rd29A啟動子上,并將該載體轉(zhuǎn)化到農(nóng)桿菌LBA4404中,通過葉盤轉(zhuǎn)化法對77個外植體進行遺傳操作,獲得了263個轉(zhuǎn)基因材料,篩選出長勢良好的轉(zhuǎn)化植株53株,移栽21株。 通過分子生物學和生理學分析方法對21株轉(zhuǎn)基因植株進行檢測和鑒定,證明了外源基因已經(jīng)整合到煙草基因組中。轉(zhuǎn)基因植株移栽后,根據(jù)鹽誘導rd29A啟動子的特點,通過鹽誘導法誘導外源FIP基因的表達,綠色熒光蛋白GFP檢測和SDS-PAGE電泳分析顯示了FIP基因得到表達,并在鹽誘導時表達增強。 本實驗結(jié)果證實了真菌免疫調(diào)節(jié)蛋白可以在煙草獲得表達,為以煙草作為生物反應器生產(chǎn)真菌免疫蛋白FIP提供了材料及技術基礎。
[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.
【學位授予單位】：東北師范大學
【學位級別】：碩士
【學位授予年份】：2008
【分類號】：R392.1

【引證文獻】

相關碩士學位論文 前1條

1 王雪飛;基因重組真菌免疫調(diào)節(jié)蛋白的篩選與功能檢測[D];上海交通大學;2012年

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本文編號：2164237