中文摘要：

通过序列比对发现猪瘟病毒的强毒株和疫苗株在毒力基因Erns中存在两个差异氨基酸,据此在猪瘟病毒标准强毒sh im en株全长感染性cDNA克隆的基础上,（1）将疫苗株的Erns基因置换sh im en株的相应片段,构建嵌合型全长cDNA克隆;（2）对Erns基因进行定点突变,使293位和311位的丝氨酸和酪氨酸分别突变为天门冬氨酰和组氨酸,构建突变型全长cDNA克隆。用SacⅡ限制性内切酶将构建的全长cDNA线性化,体外转录获得基因组RNA,利用脂质体将基因组RNA转染至PK-15细胞系,连续传代,收集3代以后培养细胞,采用RT-PCR、间接免疫荧光法进行鉴定。结果表明,所构建的全长cDNA克隆不仅具有分子水平的忠实性,而且在细胞水平表现出了感染性。该研究为后续在动物水平上进一步阐明Erns基因的功能奠定基础。

英文摘要：

Two different amino acids were found to exist in virulent E^ms gene between highly virulent and avirulent strains of CSFV by sequence comparing. To explore the importance of the two amino acids in virulence of CSFV, three full - length cDNA mutations were constructed on the base of infectious cDNA clone of CSFV shimen strain. In one mutation E^ms gene of Shimen was substituted by the corresponding region of HCLV while in the other two serine and tyrosine in site 293 and 311 were mutated into aspartoyl and histidine ,respectively. Three full- length cDNA mutations were linearized by Sac Ⅱ , transcribed in vitro and transfected into PK - 15 cells. After three passages, virual RNA and protein was detected by RT - PCR and indirect immunofluorescence assay. All results proved the fidelity and infection of these full - length cDNA mutations from molecular and cellar level. This study is a basal work for subsequent animal experiments on E^ms biological function in CSFV pathogenesis.