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| Expression of Recombinant Chicken IFN-γ Gene in Baculovirus Vector System |
| WANG Hai-Yan2 LIU Sheng-Wang1** KONG Xian-Gang1 ZHAO Zhen-Hua2 |
(1. National Laboratory of Veterinary Biotechnology, Institute of Harbin Veterinary Medicine，Chinese Academy of Agricultural Sciences, Harbin 150001, China; 2 . College of Veterinary Medicine, Inner Mongolia Agricultural University,
Huhehaote 010018, China) |
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Abstract Abstract：The cDNA of ChIFN-γ gene was subcloned into BamHⅠ site of pMelBacB under the control of baculovirus polyhedrin to construct recombinant transfer vector pMelBacB-ChIFN-γ. The recombinant transfer vector pMelBacB-ChIFN-γ was then transfected into sf9 insect cells with wild-type Bac-N-Blue linear DNA using cationic liposomes. The cultures were harvested and plaque-assayed with x-gal, and blue plaques were selected as recombinant viruses. After being purified for three times and identified by PCR, the purified viruses were named rBaculovirus-ChIFN-γ. The Sf9 cells were infected with rBaculovirus-ChIFN-γ and incubated at 28 ℃, their and cultures were harvested at 24, 48, 72, 96, 120 and 144 h post-inoculation, respectively. The lysates of the cells were analyzed by SDS-PAGE, a band with a apparent molecular weight of 19 kD was observed. To further confirm this band, the lysates of sf9 insect cells infected with rBaculovirus-ChIFN-γ were analyzed by Western blot using polyclonal antibodies against ChIFN-γ, the result showed that this band represented ChIFN-γ. It was demonstrated that about 19 kD recombinant ChIFN-γ protein with activity was expressed in sf9 insect cells.
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Received: 01 January 1900
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Corresponding Authors:
LIU Sheng-Wang
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