Construction of recombinant adenoviral vector carrying the LEDGFp52 gene by homologous recombination in bacteria and its expression in vitro
- VernacularTitle:细菌内同源重组法构建LEDGFp52基因腺病毒载体及体外表达
- Author:
Hai-Sheng, ZHAO
;
Yi, WANG
- Publication Type:Journal Article
- Keywords:
LEDGFp52 gene;
recombinant adenoviral vector;
expression in vitro
- From:
International Eye Science
2006;6(5):979-983
- CountryChina
- Language:Chinese
-
Abstract:
AIM: To construct recombinant adenoviral vector carrying the LEDGFp52 gene by homologous recombination in bacteria and to detect its expression in vitro.METHODS: The LEDGFp52 gene was cloned to adenoviral shuttle plasmid pAdTrack-CMV. Then, the resultant pAdTrack-CMV-LEDGFp52 was cotransfected into BJ5 183 bacteria with the adenoviral backbone plasmid pAdeasy-1. The adenoviral plasmid carrying LEDGFp52 was generated with homologous recombination in bacteria, and the adenoviruses were produced in 293 cells. These 293 cells were then infected with adenoviruses, and the expression of LEDGFp52 was detected by CPE (cytopathic effect) and western blot.RESULTS: The titer of Ad-LEDGFp52 adenoviruses was up to 5×1012 pfu/L after proliferation in 293 cells. LEDGFp52 was expressed efficiently in 293 cells after infection.CONCLUSION: The recombinant adenoviruses vector expressing LEDGFp52 was constructed successfully and can be used in further gene transfection experiments.
