为构建一种重组乙型肝炎病毒(hepatitis B virus, HBV)复制子模型，使其能够在病毒感染的细胞中表达可视化报告基因蛋白，本研究删除HBV基因组核心蛋白(HBV core, HBc)编码区部分序列，构建HBV1.1-ΔHBc113复制子载体。利用内含肽(intein)介导蛋白拼接的特性，选取加强绿色荧光蛋白(enhanced green fluorescent protein, EGFP)和超级折叠绿色荧光蛋白(super folder green fluorescent protein, sfGFP)作为外显肽，建立EGFP^N1-8/EGFP^C9-11和sfGFP^N1-10/sfGFP^C11蛋白分裂系统。基于ΔHBc113载体，分别构建EGFP^C9-11和sfGFP^C11重组HBV复制子；应用DNA印迹法(Southern blotting)验证两种rHBV载体的细胞内复制能力。结果显示，构建的HBV1.1-ΔHBc113复制子载体能够在HBc反式互补条件下在转染细胞中形成病毒复制。构建的EGFP^C9-11和sfGFP^C11重组HBV复制子能够支持HBc互补条件下的细胞内病毒复制，并产生子代病毒颗粒；HBV重组复制子表达的EGFP^C9-11或sfGFP^C11蛋白在共表达氮端蛋白EGFP^N/sfGFP^N细胞中，通过intein介导的蛋白质高效拼接，能够形成完整的、有功能的GFP。结果表明，构建的荧光蛋白重组HBV复制子系统能够为HBV高通量药物筛选和HBV易感性研究提供实验工具，具有重要的病毒学意义和应用前景。

In the present study, we developed a recombinant hepatitis B virus (rHBV) replicon in order to visualize virus-infected living cells via reporter gene expression. We first constructed an HBV replicon vector HBV1.1-ΔHBc113, with partial deletion of the sequence encoding HBV core (HBc). The vector demonstrated a competent viral DNA replication in transfected cells, only if HBc was provided in trans. However, the replication efficiency of ΔHBc113 vector was greatly impaired by insertion of a large foreign DNA fragment. Taking advantage of the property of intein-mediated protein splicing, we chose enhanced-green fluorescent protein (EGFP) and super folder GFP (sfGFP) as exteins, and developed EGFP^N1-8/EGFP^C9-11 and sfGFP^N1-10/sfGFP^C11 split system, respectively. We further constructed EGFP^C9-11 and sfGFP^C11 rHBV replicons based on the ΔHBc113 vector, the latter of which was proved to be competent for HBc-rescued, functional intracellular replication, and produced progeny virions in the culture medium. In cells co-expressing EGFP^N or sfGFP^N, rHBV replicon-expressed EGFP^C9-11 or sfGFP^C11 was capable of working with their counterparts, respectively, forming intact and functional GFP through intein-mediated protein splicing. We thus successfully developed a fluorescent rHBV replicon system, which can be not only used for HBV infection mechanism study, but also have an extensive application prospect for high-throughput antiviral screening.