Construction and expression of virusª²like particles containing SARS CoV RNA

    LI Zhenª²Yong, JING Jianª²Zhou, LIU Mingª²Xia, SHAO Daª²Xiao, FENG Yanª²Ming, LI Zhiª²Tao, SONG Guoª²Ying, WANG Qiuª²Qi, WANG Yunª²Long, QU Lingª²Bo, ZHAO Yuª²Fen

    1Key Laboratory of Chemobiology, School of Chemistry, Zhengzhou University, Zhengzhou 450052, China, 2Sinoª²American Biotechnology Company, Luoyang 471003, China, 3Henan Provincial Research Centre of Biotechnology, Zhengzhou 450056, China

    ¡¾Abstract¡¿ AIM: To construct and express RNaseª²resistant virusª²like particles containing the RNA fragments of SARS CoV. METHODS: The primers of MS2 assembly protein gene, coat protein gene and the fragment of SARS Cov RNA polymerase gene were synthesized according to their sequences from Genbank data and were used to amplify their cDNAs by RTª²PCR. These cDNAs were purified with SABC¬ðs ResincolumTM system for DNA purification from agarose gel. Expression vector pTrc99a and their cDNA fragments were ligated together with T4 DNA ligase after digestion with restriction enzymes. The prokaryotic expression was obtained by the transformation of the recombinant plasmid constructed above into E.coli JM109. The obtained virusª²like particles were purified and tested by quantitative analysis, RTª²PCR and stable experiment. RESULTS: The RNaseª²resistant virusª²like particles containing the RNA fragments of SARS CoV was successfully constructed and remained stable for 30 days at 37¡æ. CONCLUSION: The newª²type virusª²like particles can be conveniently used as positive control of SARS clinical nucleic diagnostic reagents. ......