摘要
本实验对鸡痘病毒２８２Ｅ４株基因组ＰｓｔⅠＨｉｎｄⅢ或ＢａｍＨⅠ片段进行了克隆、鉴定，并对部分克隆进行酶切分析，在此基础上对其中６个克隆插入Ｐ１１Ｐ７．５－ＬａｃＺ报告基因盒，构建了含报告基因的重组质粒。用其中的三个重组质粒以磷酸钙法共转染鸡胚成纤维细胞（ＣＥＦ），其中有２个产生了蓝色空斑．经三次空斑纯化能稳定产生子代病毒．证明所使用ｐＦＢ１７６和ｐＦＨ１３３为病毒复制非必需片段，分别是２．９ｋｂＢａｍＨⅠ片段和４．７ｋｂ的ＨｉｎｄⅢ片段，并绘出了它们的物理图谱．
Abstract In this paper we describe the random selection and identification of two non-seeentlal regions for viral replication of fowlpox virus 282E4 strain, which was designed as pFB176, a 2. gkb BamH I fragment. and pFH133, a 4. 7kb Hind Ⅲ fragment. Eighteen cloned FPV genomic fragment(2-10kb) of FPV 282E4 strain subjected to restriction analy,sis six LacZ transfer vectors were then constructed by insertinS the P11P7. 5-LacZ gene cassette into the appropriate RE site of cloned FPV DNA. Three LacZ transfer vectors, .pFB176Z, pFH133Z and pPB175Z, were used to transfect the FPV-infected CEP cells, and two of them, pFB176Z and pPH133Z, generated recombinant blue plaques. Purification of these blue-stained plaques was attempted and all of them were successfully purified no wildtype FPV plaques reappered during subsequent passaging. pPB176 and PFH133 are non-essential for FPV replication on CEP cells and would be the candidates for the construction of FPV-based expressingvectors.Department of Veterinary Sciences,Jiangsu Agricullfura Collegl
基金
自然科学基金农业部"八五"攻关项目
