EB virus gp85 N-terminal fragment of the prokaryotic expression and preliminary identification of.docVIP

 PAGE \* MERGEFORMAT 16 EB virus gp85 N-terminal fragment of the prokaryotic expression and preliminary identification of Author: Tu Yu-east were water-Qiu Xiang Lian Cheng Feng Feng-Hua Lan Zhong-yong [Keywords:] EB virus;, gp85, N-terminal fragment;, expression;, immunogenicity Prokaryotic expression and characterization of Nterminal truncated glycoprotein gp85 of EpsteinBarr virus [Abstract] AIM: To construct a prokaryotic recombinant vector of EpsteinBarr virus (EBV) membrane protein gp85, to express the protein in E.coli and characterize the antigenicity of this nonglycosylated protein. METHODS: The BXLF2 gene coding 5’terminal truncated of EBV gp85 was amplified from the EBV strain B958 cell line with specific primers. After identification by the restriction digestion with Hind III and Xho I, the PCR product was inserted into the prokaryotic expression plasmid pGEX5T and confirmed by sequencing. The constructed prokaryotic expression vector pGEX5T85N was transformed into the competent E.coli BL21. The expressed recombinant protein gp85N was purified by affinity chromatography, characterized by Western blot, and used immunize BALB / c mice. The titer of antisevum from the immunized mice was detected by ELISA. RESULTS: Sequencing analysis revealed that the obtained truncated BXLF2 gene was identical to that published in GenBank and successfully cloned into pGEX5T. SDSPAGE showed that the expressed recombinant protein was partially soluble with a relative molecular mass of 45000. ELISA results indicated that the expressed gp85N was recognized by that the antigp85 mAb and contiserum with high titer was obtained from the immunized mice. CONCLUSION: The obtained recombinant gp85N with an excellent antigenicity should provide preliminary data for characterization of the antibody produced by the immunized mice. [Keywords] EpsteinBarr virus; N terminal fragment of gp85; expression; immunogenicity [Abstract] Objective: To construct EB virus genes in prokaryot