Bioinformatics analysis and expression of a novel protein ROP48 in Toxoplasma gondii

Jian Zhou; Lin Wang; Aihua Zhou; Gang Lu; Qihang Li; Zhilin Wang; Meiyan Zhu; Huaiyu Zhou; Hua Cong; Shenyi He

doi:10.1515/ap-2016-0042

Artikel

Bioinformatics analysis and expression of a novel protein ROP48 in Toxoplasma gondii

Jian Zhou , Lin Wang , Aihua Zhou , Gang Lu , Qihang Li , Zhilin Wang , Meiyan Zhu , Huaiyu Zhou , Hua Cong und Shenyi He

Veröffentlicht/Copyright: 30. März 2016

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Aus der Zeitschrift Acta Parasitologica Band 61 Heft 2

Abstract

Toxoplasma gondii is an obligate intracellular apicomplexan parasite, and can infect warmblooded animals and humans all over the world. In the past years, ROP family genes encoding particular proteins of T. gondii had made a great contribution to toxoplasmosis. In this study, we used multiple bioinformatics approaches to predict the physical and chemical characteristics, transmembrane domain, epitope, and topological structure of the rhoptry protein 48 (ROP48). The results indicated that ROP48 protein was mainly located in the membrane and had several positive linear-B cell epitopes and Th-cell epitopes, which suggested that ROP48 is a potential DNA vaccine candidate against toxoplasmosis. Then the PCR product amplified from the ROP48 cDNA was inserted into a pEASY-T1 vector to build a recombinant cloning plasmid. After sequencing, ROP48 was subcloned into a eukaryotic expression plasmid pEGFP-C1 to obtain pEGFP-C1-ROP48 (pROP48). After identification by PCR and restriction enzyme digestion, the recombinant plasmid pROP48 was transfected into HEK 293-T cell and identified by RT-PCR. The results showed that the eukaryotic expression plasmid pROP48 was constructed and transfected to the cells of HEK 293-T successfully. Western blotting showed that the expressed proteins can be recognized by anti-STAg mouse sera.

Keywords: Toxoplasma gondii; ROP48; Toxoplasmosis; bioinformatics analysis; cloning; expression

Acknowledgments

This work was supported, in part, by grants from the National Natural Science Foundation of China (Grant Nos. 81071373 and 81271857), the State Key Laboratory of Veterinary Etiological Biology (Grant No. SKLVEB2011KFKT005) and the Shandong Provincial Natural Science Foundation (Grant No. ZR2009CM079). JZ, SYH, AHZ, HYZ, HC and QLZ was supported by the National Natural Science Foundation of China (Grant Nos. 81071373 and 81271857). JZ, GL, LW, YLH, JJG, PXS and HQY was supported by the Shandong Provincial Natural Science Foundation (Grant No. ZR2009CM079).

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Received: 2015-9-2

Revised: 2015-10-10

Accepted: 2015-12-10

Published Online: 2016-3-30

Published in Print: 2016-6-1

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Schlagwörter für diesen Artikel

Toxoplasma gondii; ROP48; Toxoplasmosis; bioinformatics analysis; cloning; expression