金黄色葡萄球菌肠毒素B的可溶性表达及单链抗体库的构建-运动人体科学专业论文.docxVIP

并呈现于噬菌体表面，构成表型与基因型直接相连的随机多肽、蛋白、抗体库。这 并呈现于噬菌体表面，构成表型与基因型直接相连的随机多肽、蛋白、抗体库。这 种外源多肽、蛋白、抗体可以识别相应的结合分子而被筛选出来。由于其技术上 的优越性；易于扩增和选择；筛选得到的氨基酸序列可以通过DNA测序确定。因而 得到广泛应用和迅速发展。本研究用本研究主要通过PCR，克隆SEB基因到表达 载体P““”中，引入大肠杆菌BL21中，诱导表达，利用SDS．PAGE，Western检测和鉴 定．表达出的SEB，经纯化后的SEB免疫兔子，取脾，提取总RNA，做RT—PCR， 以cDAN为摸板，扩增出vH，vL片段，用柔性linker连接后，连接到噬菌粒PoANlAB58 中，建立抗体库，本研究通过改构的噬菌粒成功的构建了单链抗体库，为筛库和 SEB中毒检测，治疗等奠定了基础。结果获得了良好的可溶性表达，纯化产量为 150mg／L。纯化产物免疫兔子后，经RT-PCR和三轮PCR成功的构建了单链抗体库， 库容达10“，可用于下一步工作。 关键词金黄色葡萄球菌肠毒素B可溶性表达单链抗体噬菌体展示 Solube Solube express of SEB and construct SEB rabbit single—chain Fv library Zbang Chongxu Abstract：The study of staphylococcal enterotoxin B(SEB)have been stopped since it was discovered in 1954，SEB is common of food poisoning and toxic shock．SEB make Rhesus monkeys vomit，this mechanism is relate to stimulation of vomit of pneumogastric and sympathetic nerve，it also conduce diarrhea,excessive and frequent evacuation of watery feces，usually indicating gastrointestinal distress disorder,even shock and death．SEB is also important member of the superantigen family，which number of pathological effects in the human，as well susceptible animals．With the development of anti—tumor medicines，more attention has focused it．As superanfigen，SEB possesses the ability bind to major histocompatibil时complex class II molecules and be recognized by T cells bearing certain T·cell receptor(TCR)V beta alleles． Superantigen—mediated T cell activation requires the participation of antigen． presenting ceils(APC)．Once superantigen has bound class II MHC molecules the surface of APC，it then interact with the T eelI receptor induce T eelI activation Superantigen-mediated T lymphocyte activation，along with its consequent cytokifie production is thought to be the basis for the pathophysiology of conditions such toxic shock syndrome and possibly rheumatoid arthritis．SEB Can activate T-lymphocytes，include the release of lymphokine and the lysis of tumor cells． The output of SEB from native staphylococcal is lo