猪圆环病毒2型ORF2与T细胞表位基因重组腺病毒表达载体的构建及鉴定

Abstract

Abstract: This study was aimed to obtain expression of fusion gene （ORF2-TCE） of porcine circovirus type 2 (PCV2) in adenoviral expression system and develop PCV2 subunit vaccine. The target genes ORF2 and T cell epitope (TCE) were obtained by PCR with pMD18-T-ORF2 and pMD18-T-TCE as templates, respectively. The fusion gene ORF2-TCE was successfully obtained by overlap extension PCR method with the ORF2-linker and linker-TCE fragments as templates. The fusion gene was cloned into the transfer vector pShuttle-CMV. The PmeⅠ-linearized plasmid vector pShuttle-CMV-ORF2-TCE was transformed into Escherichia coli bacteria competent cell BJ5183, containing the pAdEasy-1 vector by electroporation. The recombinant plasmid named pAd-ORF2-TCE was obtained and identified by PCR and PacⅠenzyme digestion. To produce the recombinant adenovirus rAd-ORF2-TCE, AD293 cells were transfected with PacⅠ-linearized plasmid pAd-ORF2-TCE. The recombinant virus named rAd-ORF2-TCE were obtained and purified through vital plaque by three times and the viral titer was 10^12.3 TCID₅₀/mL. The expression of the fusion protein was verified by Western blotting and indirect immunofluorescence assay (IFA).

Key words: porcine circovirus type 2; ORF2; T cell epitope; adenoviral expression system

MO Yong-zheng, LUO Man-lin, CHEN Rui-ai. Construction and Identification of Recombinant Adenovirus Expression Vector Carrying the Fusion Genes ORF2 and T Lymphocyte Epitopes of Porcine Circovirus Type 2[J]. .

References

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Construction and Identification of Recombinant Adenovirus Expression Vector Carrying the Fusion Genes ORF2 and T Lymphocyte Epitopes of Porcine Circovirus Type 2

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