含有EB病毒相关抗原的重组蛋白疫苗的构建和表达-生物化学与分子生物学专业论文.docxVIP

台有EB病毒掴关抗琢蛉重缀蛋自疫苗麓梅建和袅这 台有EB病毒掴关抗琢蛉重缀蛋自疫苗麓梅建和袅这 摘要 摘要 EB病毒(Epstein—BanVirus，EBV)感染与多种上皮细胞和淋熙系统恶性肿瘤密 切相关，如鼻咽癌(Nasopharyngeal Carcinoma，NPC)、布氏淋融瘤(BurkN’S Lymphoma， BL)、何杰金病(Hodgkin’S Disease，HD)以及免痰缺陷病人的恶慢淋巴瘤辞。在彝 疆瘗孛，EBV仅设逡撵毪圭|羹穗定表达见耱漤铰期懿棱撬原积貘蛋白：梭撬骧l(nuclear antigen l，EBNAl)、潜伏膜蛋白l(1atent membrane protein 1，LMP t)和潜伏膜蛋白 2(1atent membrane protein 2，LMP2)。然而LMPl能使人B细胞和上皮细胞恶性转化。 因此本文将LMP2(以下简写为P2)和EBNAl(以下简写为E1)作为研究基于细胞 免疫豹奏强癣治疗瞧豢自疫蘧翡靶撬藤。考薅蟹全长熬LMP2蛋自哥麓难畈袭达，豫 了克隆含有全长LMP2编碱序列的簇因片段(LMP2)之外，还制豁了亚克隆基因片 段(P2。251、P2。254和P2．345)。对于EBNAl，绕开蛋白中的Gly-Ala重复序列，分 别克隆基于鬣白N束端和C末端编诵序列的基因片段(El一169和El*564)。此步卜，淹 了裂1鬻炭瘗番豢致死鬻子LFrg△3冀羧弱投递功姥，涛上述撬覆基瓣片段与LFrd△3 融合，剥用pETl5b袭达质糙在大肠杼菌宿主BL21(DE3)pLysS和BL21(DE3)Codon Plus中进行栽达尝试。获得了LFn P2*251、△3 P2—251、LFn E1—169、△3 E1—169、LFn E1．564和△3 E1．564等重组融合蛋岛的表达，并迸幸亍了g、Jresterrl—Blotting检测和蛋白鹋 小援模楚亿，为逶一步静疫落骚究葵定了基番窭。 关键词：EB病毒；LMP2．EBNAl；LFrd△3：pETl5b 含有EB稍毒柜美抗穗驰重缀餐白疰营瓣鞫建帮衰遮Abstract 含有EB稍毒柜美抗穗驰重缀餐白疰营瓣鞫建帮衰遮 Abstract Epstein·Baxr virus(EBV)infection is closely related with several malignant epithelial and lymphatic tumors，such as Nasopharyngeal Carcinoma(NPC)，Bttrkitt’s Lymphoma (BL)，Hodgkin’s Disease(HD)and malignant tymphomas in immtme deficient patients．In NPC，EBV only selectively and steadily expresses a few latent nuclear antigens and membrane proteins：EBNAl(nuclear antigen 1)，LMPl(1atent membrane protein 1)and LMP2(1atent membrane protein 2)．However,LMP 1 can transform human lymphoeytes and epithelia into malignancy．Therefore，EBNAl峭1)and LMP2(p2)were chosen to be the target antigens for the development of a therapeutic protein vaccine against NPC based on 也e cell．mediated immunity．Considering the possible difficulties of expressing the full一length LMP2 protein,the sub-clones(P2-251，P2—254 and P2-345)were prepared in addition to the clone of the full．1ength LMP2 gene．For EBNAl，the repeat sequence of Gly—Ala in the protein was bypassed and the sub*clones(El一169 and