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Construction of Recombinant Adenovirus Expressing the B,C Antigenic Sites of
Spike Gene from Transmissible gastroenteritis virus(TGEV) and Its Immunization |
| TANG Fang1 ; HE Kong-wang ; HOU Ji-bo ; JIANG Ping |
| Institute of Veterinary Medicine, Jiangsu Academy of Agricultural Sciences, Nanjing 210014, China |
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Abstract The DNA fragment of 1.0 kb containing the B,C antigenic sites of the Transmissible gastroenteritis virus (TGEV) spike (S) gene was amplified by RT-PCR and cloned into the transfer vector pShuttle-CMV. The recombinant plasmid was co-transformed into Escherichia coli strain BJ5183 together with the Adenovirus (Ad) backbone vector pAdEasy-1 to generate the recombinant Ad plasmid by homologous recombination. A recombinant replication-defective human Adenovirus serotype 5 (Ad-5) containing 5' gene of TGEV S protein (rAd-TGEV-S) was generated by transfection and plaque purification in HEK293 cells. The transcription and expression of the target gene fragment was confirmed by RT-PCR and indirect immunofluorescent assay (IFA) subsequently. The recombinant virus rAd-TGEV-S was passaged by 9 times in HEK293 cells and the titer was stable with the average of 107.7TCID50. The immunization of mice with rAd-TGEV-S showed that TGEV-specific IgG and IgA were monitored.
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Received: 14 May 2008
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Corresponding Authors:
HE Kong-wang
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