用lac基因筛选表达狂犬病毒糖蛋白的重组痘苗病毒

为了研制基因工程狂犬病疫苗,我国于1991年首次报道了在痘苗病毒天坛株中表达狂犬病毒糖蛋白,但报道中重组病毒的选择是先经人骨髓瘤细胞(TK-143)在诱变剂5-溴脱氧尿苷(BrudR)作用下通过标记拯救技术筛选出携带有同源基因的重组病毒,然后再利用重组病毒中携带的Lac基因为选择标记,通过噬斑纯化获得重组病毒,用这种选择方式获得的重组病毒,经过了TK-143细胞和BrudR,因此不宜发展成疫苗,本研究探索不经过TK-143细胞和BrudR,仅利用Lac基因为选择标记,直接在鸡胚细胞上通过噬斑纯化获得重组病毒,现将研究结果报道如下。

Isolation of Vaccinia Virus Recombinants Expressing Rabies Virus Glycoprotein by Lac Gene

The expression plasmid pWS-4 containing Rabies virus glycoprotein gene was recombinaed with vaccinia viruses in the presence of Lipofectin~(TM) Reagent by homologous recombination technique in chicken embryonic cells. The Lac gene on the recombinant was used as a selected marker for isolation of TK recombinants with blue plaque on cells. Dot blot showed that the rabies virus glycoprotein gene was inserted into vaccinia genome. The recombinant was responsed specifically with McAb of rabies virus ERA strain by indirect immun of fluorescence test. Mice immunlized with recombinant produce ralization antibodies against rabies virus and were protected from the lethal challenge of rabies virus CVS strain.