1．3倍体HBV YIDD变异株真核表达载体构建及鉴定

构建HBV YIDD拉米夫定耐药株1．3倍全基因真核表达载体，为进一步探讨乙肝病毒变异株的生物学特性及筛选抗病毒药物奠定基础。参考GenBankHBV序列设计并合成一系列引物，以临床证实为拉米夫定耐药的病人HBV DNA为模板，通过PCR扩增得到HBV全基因组并克隆至pGEM—T Easy载体中，经测序证实聚合酶基因存在YIDD变异，然后以该病人的HBV全基因组为模板构建1．3倍全基因HBV—YIDD变异真核表达载体pcDNA3．1（＋）-1．3HBV。通过PCR扩增，酶切及测序证明pcDNA3．1（＋）-1．3HBV表达载体构建成功，该表达载体的构建为后期建立稳定表达HBV—YIDD变异的细胞模型提供材料。

Construction and Identification of Eukaryotic Expression Vector of 1.3 Ploid HBV YIDD Mutant Strain

In order to further investigate the biological characters of HBV mutant strain and to lay a foundation for screening antiviral medicine, a 1. 3 ploid omni-genetic eukaryotic expression vector of HBV YIDD of lamivudine drug resistant strain was constructed. Referring the sequence of HBV through GenBank, design and synthesize a series of primers, using HBV DNA from patient that was lamivudine drug resistant as a template, and then amplified with PCR and obtained HBV omni-genome and cloned into pGEM-T Easy vector, after proving there was a YIDD mutation in the polymerase gene through sequencing, finally using the patient＇s HBV omni-genome as template the 1.3 ploid omni- genetic HBV YIDD mutant eukaryotic expression vector pcDNA3.1 （ ＋ ） -1.3HBV was constructed. The result showed through PCR amplification, enzyme digestion and sequencing, and proved that pcDNA3.1 （ ＋ ）-1.3HBV was successfully constructed. The construction of the expression vector had provided a material for later to establish stably express HBV YIDD mutant cell model.