Cloning and Expression
of Chinese Duck Interferon-γ Gene
LONG Jian-Er, HUANG Li-Na, WANG Wen-Yi, CHENG Min-Jie, WEN
Yu-Mei, YUAN Zheng-Hong, QU Di*
( Department of Molecular Virology, Medical Center of Fudan University,
Shanghai 200032, China )
Abstract The
efficacy of cytokine therapy has been demonstrated in several viral diseases.
Interferon-γ is a cytokine that has potent antiviral property and
immunomodulatory activity. To investigate the role of IFN-γ in viral clearance
during natural infection and to define the antiviral mechanism, DHBV-infected
ducks was used as an animal model. To clone, express, and develop the method of
quantifying DuIFN-γ gene transcription and expression, DuIFN-γ
cDNA was amplified by RT-PCR from PHA stimulated duck PBMC. Recombinant plasmid
expressing DuIFN-γ was used to transfect COS-7, and the cell culture
supernatant was analyzed by CPE inhibitory assay and MTT methods to determine
the antiviral titer of IFN-γ. The GST-DuIFN-γ fusion protein was expressed in E.coli
and purified using the GST sepharose 4B. Results indicated that the supernatant
collected from COS-7 cells transfected with DuIFN-γ cDNA was able to
prevent duck fibroblasts from VSV induced CPE in a dose dependent manner. An
anti-DuIFN-γ antibody neutralized this antiviral activity.
Key words duck interferon-γ； cloning and expression
*Corresponding author： Tel, 86-21-64041900-2116； Fax, 86-21-64174578； e-mail, [email protected]