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题名:Inhibition of SARS-CoV replication cycle by small interference RNAs silencing specific SARS proteins, 7a/7b, 3a/3b and S
作者:Sara ?kerstr?m, Ali Mirazimi and Yee-Joo Tan
来源:Antiviral Research[IF=3.406], Volume 73, Issue 3, March 2007, Pages 219-227
URL :http://dx.doi.org/10.1016/j.antiviral.2006.10.008
日期:070415
摘要: Sara ?kerstr?ma, Ali Mirazimia, , and Yee-Joo Tanb

aCenter for Microbiological Preparedness, Swedish Institute for Infectious Disease Control, SE-171 82 Solna, Sweden
bInstitute of Molecular and Cell Biology, 61 Biopolis Drive, 138673 Singapore, Singapore

Received 17 February 2006; accepted 17 October 2006. Available online 7 November 2006

Summary

The severe acute respiratory syndrome coronavirus (SARS CoV) genome has 14 potential open reading frames (ORFs). The first ORF is translated from the full-length genomic mRNA while the remaining ORFs are translated from eight subgeomic RNAs (sgRNAs). In this study, we designed small interference RNAs (siRNAs) targeting sgRNA 2, 3 and 7 and tested their efficiency and specificity in silencing the protein translated from the targeted sgRNA. Our results demonstrated that siRNA 7 could inhibit sgRNA 7, which showed 19/19 nucleotides (nt) matching, and sgRNA 8, which showed 18/19 nt matching; but, it did not inhibit the full-length genomic mRNA which showed 17/19 nt matching. Overall, each of the siRNAs can inhibit the targeted sgRNA without affecting the full-length genomic mRNA or the other sgRNAs that showed mismatch of two or more nt. Thus, siRNA could be designed so as to knockdown the expression of viral protein(s) from a targeted sgRNA during viral infection, thereby allowing the contribution of individual viral proteins to viral infection to be delineated. When Vero E6 cells expressing siRNA 2, 3 or 7 were infected with SARS-CoV, a significant reduction in the yield of progeny virus was observed. Indirect immunofluorescence assays showed that in the infected cells expressing each of the siRNAs, there was aspecific silencing of S, 3a and 7a, respectively, but the expression of nucleocapsid protein was not affected. Thus, our data suggests that the accessory proteins, i.e. 3a and 7a, could play an important role during the replication cycle of the SARS-CoV. 29.

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