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题名:Derivation of a novel SARSCcoronavirus replicon cell line and its application for anti-SARS drug screening ? ARTICLE
作者:Feng Ge, Yonghu Luo, Pei Xiong Liew and Eugene Hung
来源:Virology[IF=3.08], Volume 360, Issue 1, 30 March 2007, Pages 150-158
URL :http://dx.doi.org/10.1016/j.virol.2006.10.016
日期:070415
摘要: Feng Gea, , , Yonghu Luob, Pei Xiong Liewc and Eugene Hunga

aDepartment of Microbiology, Yong Loo Lin School of Medicine, National University of Singapore, Block MD4, 5 Science Drive 2 Singapore 117597, Republic of Singapore
bGuangdong Institute of Microbiology, 100 Xianlie Central Road, Guangzhou, Guangdong, 510070, PR China
cDepartment of Haematology, Singapore General Hospital, Singapore Health Research Facilities, 7 Hospital Drive, Block A, 02-05, Singapore 169611, Republic of Singapore

Received 9 August 2006; revised 30 August 2006; accepted 1 October 2006. Available online 13 November 2006.

Summary

The severe acute respiratory syndrome (SARS) outbreak in 2002, which had a high morbidity rate and caused worldwide alarm, remains untreated today even though SARS was eventually isolated and controlled. Development and high-throughput screening of efficacious drugs is therefore critical. However, currently there remains a lack of such a safe system. Here, the generation and characterization of the first selectable, SARSCcoronavirus (SARSCCoV)-based replicon cell line which can be used for screening is described. Partial SARSCCoV cDNAs and antibiotic resistance/reporter gene DNA were generated and assembled in vitro to produce the replicon transcription template, which was then transcribed in vitro to generate the replicon RNA. The latter was introduced into a mammalian cell line and the transfected cells were selected for by antibiotic application. For the antibiotic-resistant cell lines thus generated, the expression of reporter gene was ensured by continued monitoring using fluorescent microscopy and flow cytometry. The suitability of this replicon cell line in drug screening was demonstrated by testing the inhibitory effect of several existing drugs and the results demonstrate that the SARSCCoV replicon cell lines provide a safe tool for the identification of SARSCCoV replicase inhibitors. The replicon cell lines thus developed can be applied to high-throughput screening for anti-SARS drugs without the need to grow infectious SARSCCoV. 13.

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