CAJ | 학술논문

目的构建含有A亚型人呼吸道合胞病毒(Human Respiratory Syncytial Virus,RSV)融合糖蛋白(Fusion glycoprotein,F)基因的非复制型第一代重组腺病毒(First generation odenovirns vector,FGAd),并研究F基因在重组腺病毒中的表达.方法利用限制性内切酶Xho Ⅰ和HindⅢ从质粒pGEM3zf-F中切下目的基因F,克隆至穿梭质粒pShuttle-CMV,再与pAdeaRy-1在大肠埃希菌BJ5183中进行同源重组,鉴定正确后,用脂质体法转染293细胞,Western Blot鉴定目的基因表达.结果获得了表达RSV F基因的非复制型重组腺病毒FGAd/F,Western Blot检测到F基因的表达.结论获得一株可表达A亚型BSV F的非复制型重组腺病毒FGAd/F,可用于体内研究观察其免疫效果及免疫保护作用.
목적 구건함유A아형인호흡도합포병독(Human Respiratory Syncytial Virus,RSV)융합당단백(Fusion glycoprotein,F)기인적비복제형제일대중조선병독(First generation odenovirns vector,FGAd),병연구F기인재중조선병독중적표체.방법 이용한제성내절매Xho Ⅰ화HindⅢ종질립pGEM3zf-F중절하목적 기인F,극륭지천사질립pShuttle-CMV,재여pAdeaRy-1재대장애희균BJ5183중진행동원중조,감정정학후,용지질체법전염293세포,Western Blot감정목적 기인표체.결과 획득료표체RSV F기인적비복제형중조선병독FGAd/F,Western Blot검측도F기인적표체.결론 획득일주가표체A아형BSV F적비복제형중조선병독FGAd/F,가용우체내연구관찰기면역효과급면역보호작용.
Objective A strain of replication deficient recombinant adenvirus encoding fusion glycoprotein (F) of subgroup A human respiratory syncytial virus (RSV) was constructed and the expression of F was identified. Methods The F gone was obtained from pGEM3zf-F with Xho Ⅰ and Hind Ⅲ ,cloned into adenovirase shuttle vector pShutth-CMV,and then the resulting pShutth-CMV/F was transformed into E. coli BJS183/p with pAdeasy-1 to produce pre-adenoviral plasmid encoding F by homologous recombination. This resultant plasmid was linearized by digestion with Pac Ⅰ and transfected into 293 packaging cells to generate FGAd-F. Finally,the expression of F protein was identified by Western Blot analysis. Results FGAd/F was successfully constructed,and the expression of RSV F protein was identified by Western Blot. Conclusion We have obtained a strain of rephcation-defective adenovirus FGAd/F encoding RSV F protein,which can be used further to investigate its protective efficacy against RSV infection in vivo.