本文關(guān)鍵詞：伯氏瘧原蟲類巨噬細(xì)胞移動(dòng)抑制因子的表達(dá)、純化及其在紅內(nèi)期表達(dá)的檢測(cè)　出處：《第四軍醫(yī)大學(xué)》2006年碩士論文　論文類型：學(xué)位論文

  更多相關(guān)文章： 伯氏瘧原蟲 巨噬細(xì)胞移動(dòng)抑制因子 基因表達(dá) 多克隆抗體 免疫印跡

【摘要】：目的：瘧疾是世界上對(duì)人類危害最嚴(yán)重的傳染病之一。目前對(duì)瘧疾的防治效果不理想，，其原因在于對(duì)瘧原蟲的生物學(xué)特性及其與宿主之間的相互作用缺乏了解。瘧原蟲基因組計(jì)劃的實(shí)施，使人們發(fā)現(xiàn)大量未知功能的基因。對(duì)未知功能基因的研究不僅給深入了解瘧原蟲生物學(xué)特性提供巨大的幫助，而且為尋找新的藥物作用靶點(diǎn)和疫苗的研制提供新的思路。巨噬細(xì)胞移動(dòng)抑制因子(MIF)是較早發(fā)現(xiàn)的廣泛存在于人及其他哺乳動(dòng)物的細(xì)胞因子，可廣泛參與哺乳動(dòng)物的多種生物反應(yīng)，特別是免疫功能的調(diào)節(jié)。瘧原蟲也具有MIF的同源基因。對(duì)瘧原蟲類MIF的研究可能會(huì)給深入了解瘧原蟲的生物學(xué)特性提供幫助。為研究瘧原蟲MIF生物學(xué)功能，本研究擬對(duì)伯氏瘧原蟲類巨噬細(xì)胞移動(dòng)抑制因子(PbMIF)進(jìn)行克隆和表達(dá)，獲取純化的可溶性蛋白，制備小鼠抗PbMIF多克隆抗體，并檢測(cè)PbMIF在伯氏瘧原蟲紅內(nèi)期的表達(dá)。 方法：根據(jù)GenBank中PbMIF mRNA的預(yù)測(cè)序列，設(shè)計(jì)特異引物，采用RT-PCR方法從伯氏瘧原蟲ANKA株紅內(nèi)期RNA擴(kuò)增獲得PbMIF基因。將PbMIF基因與T載體連接并測(cè)序，利用NCBI中Blast程序分析測(cè)序結(jié)果。陽(yáng)性T／A克隆質(zhì)粒經(jīng)BamH Ⅰ和Xho Ⅰ雙酶切后，將目的片段克隆至原核表達(dá)載體pET28a，并轉(zhuǎn)化大腸埃希菌BL21(DE3)，收集經(jīng)IPTG誘導(dǎo)表達(dá)的
[Abstract]:Objective: malaria is one of the most serious infectious diseases in the world. This is due to the lack of understanding of the biological characteristics of Plasmodium and its interaction with the host, and the implementation of the Plasmodium Genome Project. The study of unknown functional genes not only provides a great help for further understanding the biological characteristics of Plasmodium falciparum. And it provides a new way to find new drug target and vaccine. Macrophage migration inhibitory factor (MIF) is widely found in human and other mammalian cytokines. It can be widely involved in a variety of mammalian biological reactions. In particular, the regulation of immune function. Plasmodium also has the homologous gene of MIF. The study on MIF of Plasmodium may be helpful to understand the biological characteristics of Plasmodium, and to study MIF biology of Plasmodium. Learning function. In this study, Plasmodium berghei macrophage migration inhibitor PbMIF was cloned and expressed to obtain the purified soluble protein and to prepare mouse anti-PbMIF polyclonal antibody. The expression of PbMIF in erythrocyte of Plasmodium berghei was detected. Methods: according to the predicted sequence of PbMIF mRNA in GenBank, specific primers were designed. The PbMIF gene was amplified by RT-PCR from the erythroid RNA of Plasmodium berghei ANKA strain. The PbMIF gene was ligated with T vector and sequenced. The positive T / A clone plasmid was digested by BamH 鈪

本文編號(hào)：1441660