[Construction of recombinant vaccinia virus expressing HSV-2 gD gene as live recombinant vaccine strain].
We had reported that the recombinant vaccinia virus expressing glycoprotein D of herpes simplex virus type 2 (HSV-2 gD) protected mice against lethal HSV-2 challenge. Following the succeed in animal model, we continue the research to construct the recombinant vaccinia virus expressing HSV-2 gD gene as live recombinant vaccine strain in strict accordance with the guideline for human vaccine research. A PCR-modified HSV-2 gD gene was inserted into the plasmid pJSB1175, under the control of P7.5K early/late promoter of vaccinia virus. The recombinant plasmid was used, with lipofectin reagent, to transfect 2BS cells which had been infected by wild type of TK+ vaccinia virus (Tian Tan 761 strain). The recombinant vaccinia virus harboring HSV-2 gD gene was selected out by using in situ hybridization employing 32P-1a-belled HSV-2 gD fragment as probe, together with three cycles of plaque purification. Dot and Southern blot confirmed HSV-2 gD gene had been integrated into the TK region of vaccinia virus genome, as expected. Indirect immunofluorescent assay using anti-HSV-2 gD monoclonal antibody showed HSV-2 gD was expressed effectively in the recombinant virus infected cells.
Yan
H
H
Institute of Virology, Chinese Academy of Preventive Medicine, Beijing 100052.
Rong
M
M
Ruan
L
L
Hou
Y
Y
He
N
N
Zhu
J
J
D000818:Animals; D015139:Blotting, Southern; D002478:Cells, Cultured; D002642:Chick Embryo; D018258:Herpesvirus 2, Human / Q000235:genetics* / Q000276:immunology; D006801:Humans; D017403:In Situ Hybridization; D010957:Plasmids / Q000235:genetics; D011995:Recombination, Genetic; D014614:Vaccines, Synthetic / Q000235:genetics* / Q000276:immunology; D014616:Vaccinia virus / Q000235:genetics* / Q000276:immunology
D014614:Vaccines, Synthetic