猪圆环病毒2型DY株ORF2基因在Sf9昆虫细胞中的表达

Abstract

Abstract: The ORF2 gene of porcine circovirus type 2(PCV2) including complete open reading frame(ORF)was amplified by PCR,and it was cloned into the pFastBac1^TM vector of Bac-to-Bac baculovirus expression systems to construct the recombinant transfer vector pFast-ORF2.The transfer vector was transformed into DH10Bac competent cell which contained the BmNPV bacmid. A recombinant shuttle vector which contained ORF2 gene was constructed by site-specific transposition and the colonies containing recombinant bacmid were collected by white selection. The cultured Sf9 cells were transfected with recombinant bacmid DNA mediating with cellfectin reagent and the pure recombinant baculovirus Bacmid-ORF2 was obtained. After transfecting this transformant into Sf9 cells,Cap protein expression in Sf9 cells was detected by indirect immunofluorescent assay and Western blotting. The experimental results showed that the recombinant baculovirus was obtained and the Cap protein with a molecular weight of 31 ku was expressed in insect Sf9 cells. The recombinant protein could be recognized by PCV2 positive serum. This would lay a foundation for developing PCV2 subunit vaccine and diagnosis method.

Key words: porcine circovirus type 2; ORF2 gene; Sf9 insect cells; expression

CLC Number:

WANG Wen-xiu, SHEN Zhi-qiang, WANG Shan-hui, ZHANG Song-lin, CHI Xian-feng, XIA Xiao-jing. Expression of ORF2 Gene of Porcine Circovirus Type 2 in Sf9 Insect Cells[J]. , 2012, 39(10): 53-57.

References

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Expression of ORF2 Gene of Porcine Circovirus Type 2 in Sf9 Insect Cells

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