摘要
目的以原代培养的小鼠卵巢颗粒细胞为模型,观察人SET基因重组腺病毒载体在卵巢颗粒细胞的感染效率和基因表达效果。方法分离并培养小鼠颗粒细胞。扩增人SET基因重组腺病毒载体AdCMV-SET和对照重组腺病毒载体AdCMV,感染原代培养的小鼠颗粒细胞,通过观察绿色荧光蛋白(GFP)检测感染效率;qRT-PCR方法检测SETmRNA表达;Western blot方法检测SET蛋白的表达水平。结果获得的重组腺病毒载体体外感染小鼠颗粒细胞,24h观察到GFP表达。与AdCMV感染组相比,AdCMV-SET感染组SETmRNA表达水平显著增高(P<0.05)。与空白对照组(Mock)及感染AdCMV组相比,AdCMV-SET感染组SET蛋白水平显著增高(P<0.05)。结论构建的人SET基因重组腺病毒载体能高效感染小鼠颗粒细胞、并有效表达,对细胞活率无影响。
Objective The primary cultured granulosa cells of mice were chosen to study the efficiency and effectiveness of delivering SET gene by the SET gene recombinant adenovirus vector in reproductive-related cells.Methods SET gene recombinant adenovirus vector AdCMV-SET and the control adenovirus vector AdCMV were amplified.The primary mouse granulosa cells were infected by the recombinant adenovirus and the infection efficiency was observed through the green fluorescence protein（GFP）.SET mRNA expression was detected by qRT-PCR and SET protein was detected by Western blot.Results After infected by the recombinant adenovirus vector for 24 h,GFP expression was observed in the granulosa cells.Compared to the AdCMV-infected mouse granulosa cells,the mouse granulosa cells infection of AdCMV-SET were significantly increased（P0.05）.SET protein levels were significantly higher in the mouse granulosa cells infected by AdCMV-SET than those in both the uninfected group（Mock） and the group of AdCMV（P0.05）.Conclusion The recombinant adenovirus vector AdCMV-SET is successfully constructed and effectively expresses SET mRNA and SET protein in mouse granulosa cells,which has no effect on the activity of the mouse granulosa cells.
出处
《江苏医药》
CAS
CSCD
北大核心
2010年第16期1916-1919,F0003,共5页
Jiangsu Medical Journal
基金
国家教育部创新团队课题(IRT0631)
国家自然科学基金(30800394
30801236)
