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| Rescue of the Minigenome of Human Respiratory Syncytial Virus Based on T7 Promoter Expression System |
| YUAN Rui1,2, FU Yuan-hui2, HE Jin-sheng2, JIAO Yue-ying2, JIANG Gui-yun1,2, ZHANG Mei1, PENG Xiang-lei2, KAN Xue-tong 1,2 |
1. Department of Immunology, Anhui Medical University, Hefei 230032, China;
2. College of Life Sciences and Bioengineering, Beijing Jiaotong University, Beijing 100044, China |
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Abstract Objective:To establish a T7 promoter based reverse genetics system competent for the rescue of human respiratory syncytial virus. Methods:We construct four helper plasmids of px8δT-PT1-N, px8δT-PT1-P, px8δT-PT1-M2-1 and px8δT-PT1-L encoding RSV nucleocapsid proteins, respectively, and one minigenome plasmid of pSC11-E containing open reading frame of the enhanced green flurorecent protein and cis-acting elements including RSV leader region/promoter, gene start, gene end and trailer region/antigenomic promoter. All these plasmids are under the control of T7 promoter and identified by restriction endonucleoase analysis and Western blot. The pSC11-E is rescued by RSV and the above helper plasmids in BSR T7/5 cells, respectively. Then, the fluorescence expression is observed over time with fluorescence microscopy. Results:We successfully constructed a reverse genetics system based 5 plasmids undert the control of T7 promoter and finished the rescue operation to the minigenome of RSV. Conclusion:This system can be further applied to investigate the function of RSV genome by deletion and mutation of its genes.
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Received: 25 April 2013
Published: 25 September 2013
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