Cloning, expression, and immunoreactivity analysis of antigen Eg-08002 from protoscoleces of Echinococcus granulosus

Abstract

Abstract:

Objective To clone, express and analyze the immuno-reactivity of Eg-08002, an antigen screened out from protoscolex-stage Echinococcus granulosus.Methods Based on a previous study of transcriptome sequencing at the protoscolex phase and oncosphere, the protoscolex stage specific antigen Eg-08002 was identified and amplified by PCR and cloned into vector pET28a to construct recombinant plasmid pET28a-Eg-08002, which was then transformed into Escherichia coli JM109 for expression under the induction of IPTG.The recombinant rEg-08002 protein was purified with His-tag Ni-superflow affinity chromatography, and verified by rapid non-staining polyacrylamide electrophoresis.Immunoreactivity of rEg-08002 was analyzed by Western blotting, which was then assessed with sera from 12 patients with cysitic echinococcosis and 12 healthy individuals by ELISA.Results High expression of Eg-08002 was detected in protoscolex, with a length of 612 bp.The recombinant pET28a-Eg-08002 was constructed.Results of rapid polyacrylamide gel electrophoresis and Western blotting indicated that the recombinant protein rEg-08002 was highly expressed in E.coli JM109 in a predominant form of inclusion bodies, with an M_r of 23000, and was recognized by sera from mice with cystic echinococcosis.ELISA results showed that the average A₄₅₀ values with sera from the patients and healthy individuals were 1.245 ± 0.265 and 0.469 ± 0.006, respectively, with statistical difference（P < 0.05）.Conclusion The antigen Eg-08002 was identified with high expression in the protoscolex phase with a promising immunoreactivity as revealed by Western blotting and ELISA

Key words: Echinococcus granulosus, Cloning, Expression, Immunoreactivity

CLC Number:

R383.3

Hui YANG, Yin-qi ZHAO, Zi-hua LI, Jia-qing ZHAO, Hao WANG, Ya-na WANG, Ming-xing ZHU, Wei ZHAO. Cloning, expression, and immunoreactivity analysis of antigen Eg-08002 from protoscoleces of Echinococcus granulosus[J]. CHINESE JOURNAL OF PARASITOLOGY AND PARASITIC DISEASES, 2017, 35(6): 559-562.

Figures/Tables 3

References 18

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