The preparation of recombinant adenovirus Ad-Rad50-GFP and detection of the optimal multiplicity of infection in CNE1 transfected hv Ad-Rad50-GFP.
- Author:
Ruicheng YAN
;
Jiancong HUANG
;
Ling ZHU
;
Lihong CHANG
;
Jingjia LI
;
Xifu WU
;
Jin YE
;
Gehua ZHANG
- Publication Type:Journal Article
- MeSH:
Adenoviridae;
Carcinoma;
Cell Line, Tumor;
Genetic Vectors;
Humans;
Nasopharyngeal Carcinoma;
Nasopharyngeal Neoplasms;
Transfection
- From:
Journal of Clinical Otorhinolaryngology Head and Neck Surgery
2015;29(24):2143-2146
- CountryChina
- Language:Chinese
-
Abstract:
OBJECTIVE:The optimal multiplicity of infection (MOI) of the recombinant adenovirus Ad-Rad50-GFP carrying a mutant Rad50 gene expression region on the cell growth of nasopharyngeal carcinoma and the viral amplification efficiency of CNE1 cell infected by this adenovirus were studied.
METHOD:The biological titer of Ad-Rad50-GFP was measured by end point dilution method. The impact of recombinant adenoviral vector transfection on the growth of CNE1 cells was observed by cell growth curve. Transfection efficacy of recombinant adenoviral vector was observed and calculated through fluorescence microscope. The expression f mutant Rad50 in the Ad-Rad50-GFP transfected CNE1 cells with optimal MOI was detected by Western Blot after transfection.
RESULT:The biological titer of Ad-Rad50-GFP was 1.26 x 10¹¹ pfu/ml. CNE1 cell growth was not influenced significantly as they were transfected by recombinant adenoviral vector with MOI less than 50. Transfection efficacy of recombinant adenoviral vector was most salient at 24 hours after transfection, with the high expression of mutant Rad50, and the efficiency still remained about 70% after 72 hours.
CONCLUSION:Recombinant adenoviral vector Ad-Rad50-GFP could transfect CNE1 cells as well as result in the expression of mutant Rad50 in CNE1 cells effectively. MOI = 50 was the optimal multiplicity of infection of CNE1 cells transfected by recombinant adenoviral vector Ad-Rad50-GFP.
