Recovery of infectious human parainfluenza virus type 3 from cDNA
Recovery of infectious human parainfluenza virus type 3 from cDNA
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DOI：
10.1006/viro.1997.8697
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发表时间：
1997-09-01
期刊：
影响因子：
3.7
通讯作者：
Murphy, BR
中科院分区：
文献类型：
--
作者：
Durbin, AP;Hall, SL;Murphy, BR
传染性HPIV3是由四种质粒携带的cdna在细胞内共表达产生的。这些基因分别编码了一个完整的HPIV3基因组（负义）、HPIV3核衣壳蛋白N、磷酸化蛋白P和聚合酶蛋白l。cdna编码的HPIV3基因组与构建HPIV3时使用的JS野生型（wt）菌株存在7个点突变：其中四个是作为dna衍生病毒标记的HN或L基因编码区域的沉默突变，两个被引入以产生氨基酸替代，使HN特异性单克隆中和抗体423/6识别的表位消失，其余的点突变导致HN蛋白氨基酸263位的偶然氨基酸替代。将这4个质粒转染到HEp-2细胞单层中，由重组痘苗病毒提供的T7 RNA聚合酶驱动其表达。收获的转染上清中存在的病毒滴度低(
Infectious HPIV3 was produced by the intracellular coexpression of four plasmid-borne cDNAs. These separately encoded a complete HPIV3 genome (negative-sense), the HPIV3 nucleocapsid protein N, the phosphoprotein P, and the polymerase protein L. The cDNA-encoded HPIV3 genome differed from the JS wildtype (wt) strain of HPIV3 used in its construction by seven point mutations: four of these are silent mutations in the HN or L gene coding regions that serve as markers of a cDNA-derived virus, two were introduced to create an amino acid substitution that ablates an epitope recognized by the HN-specific monoclonal neutralizing antibody 423/6, and the remaining point mutation results in an incidental amino acid substitution in the HN protein at amino acid position 263. The four plasmids were transfected into HEp-2 cell monolayers and their expression was driven by T7 RNA polymerase supplied by a vaccinia virus recombinant. The titer of virus present in the harvested transfection supernatant was low (