牛巴贝斯虫SBP2蛋白生物信息学分析及多克隆抗体制备

doi:10.16431/j.cnki.1671-7236.2025.08.033

Abstract

Abstract: 【Objective】 The experiment aimed to explore the bioinformatics characteristics of the spherical body protein 2 (SBP2) of Babesia bovis,and conduct prokaryotic expression and prepare polyclonal antibodies,providing a theoretical basis for the screening of Babesia bovis vaccines and diagnostic antigens. 【Method】 SBP2 gene of Babesia bovis was amplified and cloned. The phylogenetic tree of SBP2 protein of Babesia bovis was constructed using Mega 7.0.The phosphorylation sites and B-cell antigenic epitopes of the SBP2 protein in Babesia bovis were predicted and analyzed using bioinformatics methods such as IEDB Analysis Resource.The amino acid sequences of SBP2 protein and dense granules antigen (GRA) of Toxoplasma gondii were compared and analyzed. The prokaryotic expression vector pET-28a-SBP2 was constructed. The recombinant protein of SBP2 was induced to be expressed and purified.The reactivity of the recombinant protein of SBP2 was verified by Western blotting.Polyclonal antibodies were prepared by immunizing BALB/c mice with the recombinant protein SBP2 as the immunogen,and their titers were detected by indirect ELISA method. 【Result】 The phylogenetic tree showed that SBP2 protein of Babesia bovis in this study was most closely related to the amino acid sequence of the Thai strain (OM46855).Bioinformatics analysis revealed that SBP2 protein contained 17 B-cell antigenic epitopes and 31 phosphorylation sites,including 12 serine sites,14 threonine sites and 5 tyrosine sites.There was a strong correlation between the amino acid sequence of SBP2 protein of Babesia bovis and GRA protein of Toxoplasma gondii.The recombinant plasmid pET-28a-SBP2 was successfully constructed.The results of Western blotting showed that the molecular weight of the SBP2 recombinant protein was approximately 35 ku and it had good reactivity.The titer of the prepared polyclonal antibody was 1∶204 800. 【Conclusion】 This study deepened the understanding of SBP2 protein by predicting its biological characteristics.The SBP2 recombinant protein of Babesia bovis was successfully obtained using the prokaryotic expression system,and its polyclonal antibodies derived from mice were also acquired,laying a foundation for further research on the function of SBP2 and its role in the pathogenic mechanism of Babesia bovis.

Key words: Babesia bovis; SBP2; bioinformatics analysis; prokaryotic expression; polyclonal antibodies

CLC Number:

S852.72

FENG Xiujuan, REN Jichao, CUI Zeyun, ZHAO Xueqing, LI Jiaxin, ZHANG Yang, ZHANG Wei, BAYIN Chahan·Gailike, GUO Qingyong, LI Yongchang. Bioinformatics Analysis and Preparation of Polyclonal Antibodies Against Babesia bovis SBP2 Protein[J]. China Animal Husbandry & Veterinary Medicine, 2025, 52(8): 3847-3856.

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Bioinformatics Analysis and Preparation of Polyclonal Antibodies Against Babesia bovis SBP2 Protein

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