中文摘要：

为了探究HBVx是否累及其他组织器官,本研究构建了携带HBVx基因的慢病毒表达载体,感染14-19肝前体细胞,使其能稳定表达HBVx基因,并构建了门静脉动物模型,使肝前体细胞在小鼠体内存活并能够进一步分化,并在不同时间段分别检测小鼠各个组织中HBVx目的基因的表达情况。通过PCR扩增HBVx目的基因,使之与逆转录病毒表达载体（p Lenti-CMV-HA-3FLAG-PGK-EGFP-F2A-Puro）连接,后通过酶切、菌落PCR、测序验证质粒是否构建成功。构建成功后用携带HBVx逆转录病毒感染14-19肝前体细胞,嘌呤霉素（puromycin）连续筛选4周后进行细胞HBVx目的基因检测。同时进行门静脉动物模型构建,分别在第3天、7天、14天、30天、60天进行检测小鼠各个组织器官中的HBVx的表达情况。成功构建携带HBVx的慢病毒表达质粒、稳定携带HBVx的14-19肝前体细胞、门静脉动物模型。同时检测了各个组织器官HBVx的表达情况,HBVx可在肝脏内生长分化且可在肝脏内长时间高表达,其余组织不表达。携带HBVx的肝前体细胞可在肝内生长分化,且HBV具有嗜肝性,不累及其他脏器。构建了一个长期表达HBVx的动物模型,为本课题组后续研究HBVx导致肝癌发生发展奠定了良好基础。

英文摘要：

ha order to explore whether HBVx involves other organs, we constructed lentiviral vector carrying HBVx gene. We used the constructed vector to infect 14-19 hepatic progenitor cells to made it express HBVx gene steadily; While, we constructed the model of portal vein animals to make hepatic progenitor cells surviving and further differentiating in mice, which made it possible for us to detect the expression of HBVx target gene in each organ of mice at different times. HBVx gene was amplified by PCR and connected with the lentivirus expression vector （pLenti-CMV-HA-3FLAG-PGK-EGFP-F2A-Puro）, and then restriction enzyme digestion, colony PCR and sequencing were used to verify whether the plasmid was successfully constructed. After successful construction, retrovirus carrying HBVx was adopted to infect lentivirus 14-19 hepatic progenitor cells, and the detection of HBVx target gene was carried out after continuous selection of puromycin for 4 weeks. At the same time, we constructed portal vein animal models and detected the expression of HBVx in different organs of mice at the 3^rd, 7^th, 14^th, 30^th, and 60^th day, respectively. As a result, we successfully constructed lentivirus carrying HBVx expression plasmid; 14-19 hepatic progenitor cells stably carrying HBVx stable, and the portal vein animal models. Meanwhile, we determined the expression of HBVx in various organs of mice and found that HBVx could grow and differentiate had high expression, in the liver for a long timer while it was not expressed in other organs. Furthermore, hepatic progenitor cells carrying HBVx also could grow and differentiate in liver, and HBVx was hepatotrophic, but it did not involves other organs. The build of an animal model of long-term expression of HBVx provided a good start for the follow-up study of HBVx leading to the occurrence and development of liver cancer.