乙型肝炎病毒S基因片段的克隆及其原核表达的实验研究

Cloning Study of S Gene in Hepatitis B Virus

目的 :获得adw亚型乙肝病毒表面抗原S基因在原核中的克隆和表达 ,经家兔免疫后产生抗 -HBs抗体。方法 :从乙型肝炎病人血清中提取HBVDNA后扩增出 5 0 1bp的核苷酸片段 ,经纯化限制性内切酶 (BamHⅠ和HindⅢ )切后 ,分别与PUC19和pGEMEXTM - 1载体连接后转化入宿主菌JM 10 9中和BL2 1,重组质粒经蓝 /白斑 ,筛选、PCR鉴定、酶切鉴定 ,序列分析得到阳性重组质粒pGE -S。通过IPTG诱导在大肠杆菌BL2 1中表达出重组抗原 ,重组抗原免疫家兔后 ,用ELISA法在血清中检测到抗 -HBs的存在。结果 :HBVDNA经扩增得到 5 0 1bp的核苷酸片段 ,与质粒重组后形成的重组质粒 ,在IPTG诱导形成重组抗原 ,重组抗原两次免疫家兔后的血清抗体阳性率比较 ,差别有显著性 (χ2 =0 .2 5 ,P≤ 0 .0 5 )

Objective: To study the cloning and expression of s gene in hepatitis B virus. Method:the HBV DNA was obtained from the serum of the patients with hepatitis B and dilated to the 501bp nucleotide. And they were linked with PUC19 and pGEMEX-1 by Bam H I and Hind III and to involve in the JM109 and BL21. Anti-HBs was measured by ELISA. Results and conclusion: there was a significant difference of antibody positive rate between the 501bp nucleotide after dilating HBV DNA and PGE-S(χ 2=0.25,P≤0.05).