本實驗將聚合酵素鏈反應生成的B型肝炎病毒X基因，利用基因重組技術 成功的嵌入穀胱甘汰轉移脢基因接合系統的載體pGEX-5X-1；經由轉形作 用將重組質體植入大腸桿菌DH5a 中，利用限制酵素確定X基因嵌入載體的 方向性；選殖出能誘發表達穀胱甘汰轉移脢和X抗原組成接合蛋白質的細 菌株，並以穀胱甘汰瓊脂醣珠和接合蛋白質的親和特性，純化出分子量 為43 Kd均質的接合蛋白質；利用蛋白質切割酵素Factor Xa和接合蛋白質 作用，再純化出均質的X蛋白質，並且未來將產生B型肝炎病毒X蛋白質的 抗體，以便對X蛋白質的功能有更進一步的了解。 Hepatitis B virus X gene (HBx gene) was amplified from polymerase chain reaction. The resulting DNA was successfully inserted into the plasmid pGEX-5X-1 of Glutathione S-transferase (GST) Gene Fusion System by recombinant DNA technology. The recombinant plasmid was transformed into E. coli DH5a by transformation and the orientation of the inserted HBx gene was confirmed by restriction enzyme analysis. The cloned E. coli was induced to express fusion protein, which contained GST and the HBx antigen. Due to the affinity between GST and glutathione sepharose-4B, the fusion protein with a molecular weight of 43 KD was chemically purified to homogeneity. The purified fusion protein was further digested with Factor Xa protease and the HBx antigen was purified. The HBX antigen will be used to produce antibodies for the functional identification of the HBx protein.
