摘要
目的：构建针对乙型肝炎病毒（HBV）C区基因（2029-31及2063-65位点）的双位点核酶的真核表达载体．观察其在细胞内对HBV基因表达的抑制作用．方法：利用亚克隆技术．从质粒pGEMRzl23切下EcoRI-BamHI片段，双粘端克隆于真核质粒pBBS212中，将该质粒与P1．2Ⅱ（含HBV全序列）共转染HHCC细胞（人肝癌细胞株）．观察该核酶对HBV基因表达的抑制作用．结果：在HHCC细胞中p1．2Ⅱ可表达出HBsAg及HBeAg该核酶对HBeAg的合成抑制率达65％．结论：该双位点核酶可能通过针对HBVC区mRNA的剪切作用，阻断C区mRNA的表达．抑制了HBeAg的合成。
AIM: To construct a eukaryotic expression vectorwith dual--target ribozyme against hepatitis B virlls and to observe the inhibition of HBeAg expressed in the HHCC cell(human hepatic carcinoma cell line ) transfected by HIVgenome and dual--target ribozyme against HBV. METHODS:Using subclone technique, the dual--target ribozyme gene cutfrom pGEM-Rz 123 was ligated into the eukaryotic expression vector pBBSZ12. The recombinant plasmid pBBS212uRzand pl. 2 1 carrying genome of adr--subtype HllV were cotransfected into the HHCC cell by using lipofectamine. Theintracellular e/c antigen of HBV was detected by EI.ISA andimmunohistochemistry. RESULTS: The cotransfected cellsdid express HBV e antigen and ribozyrne RNA, and the dualtarget ribozyme inhibited the level of HlleAg expressed on cotransfected cells by up to 65%. CONCLUSION: The dualtarget ribozyme can cleave the HBV core region mRNA andinhibit HBV the expression of HBV core region gene.
出处
《第四军医大学学报》
1999年第8期645-649,共5页
Journal of the Fourth Military Medical University
