Construction of Recombinant Adenovirus Expressing the B,C Antigenic Sites of Spike Gene from Transmissible gastroenteritis virus(TGEV) and Its Immunization
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.
