從2002年末至2003年初，由新種冠狀病毒所引起的嚴重急性呼吸道症候群(SARS)的疫情，在世界各地相繼爆發，引起人們對於病毒的恐慌；面對各種病毒威脅人類的健康，研究人員急欲尋求有效的抗病毒藥物，但目前仍未發展出有效藥物。而RNA干擾技術( RNA interference，RNAi )則提供了研發抗病毒藥物新的方向，原理是細胞內的小片段干擾RNA (siRNA)會活化轉錄後基因沉默作用(Post-transcriptional gene silencing，PTGS)，使具序列同源互補特性的mRNA被降解，以阻止該基因的表現；RNA干擾技術可阻斷因病毒感染或基因缺陷等異常基因的表現，達到治療的目的。C1+ SARS 1000質體可表現SARS病毒部份尖釘蛋白與綠螢光蛋白融合mRNA；將C1+ SARS 1000質體與siRNA片段表現質體共同轉染至HeLa細胞中，使尖釘蛋白的mRNA作為siRNA片段作用的受質。之後以北方吸漬法、反轉錄PCR與定量PCR加以檢測尖釘蛋白mRNA之表現量。我們證明由質體轉染所表現的siRNA片段可以有效專一的抑制SARS病毒尖釘蛋白RNA的表現，在我們所設計的4種siRNA序列中，有一siRNA序列可使細胞內的尖釘蛋白mRNA表現量降低至20%左右。為增加RNAi表現效率及降低細胞毒性，我們改以腺病毒表現載體取代轉染試劑的方式，我們觀察到其高表現量的siRNA序列更能專一性的降低細胞內SARS病毒尖釘蛋白的mRNA。我們的研究證明了結合腺病毒表現載體和RNA干擾技術的可行性，在未來也許可應用於治療SARS或其他方面的疾病。

Serious outbreaks of severe acute respiratory syndrome (SARS), caused by the newly discovered coronavirus SARS-CoV, occurred between late 2002 and early 2003. RNA interference (RNAi) is a process by which the introduced small interfering RNA (siRNA) could bind to the homologous RNA sequence and the binding adduct is degraded by RISC complex. Recently, siRNA- induced RNA interference may provide a new approach for the therapy of pathogenic or genetic diseases. The plasmid C1+ SARS 1000, which contains the genes encoding eGFP and part of SARS-Cov S protein, was co-transfected with plasmid expressing specific siRNAs for SARS-Cov S gene in HeLa cells. Among 4 siRNA sequences, we found that one siRNA, name IIIsas, could specifically reduce the expression of spike mRNA over 80% detected by Northern bloting, reverse transcription PCR and quantitative PCR. The downregulation of SARS-S gene expression was correlated with the concentrations of the siRNA expression vector employed in a range of 4.5~18 □g plasmid DNA. The siRNA was delivered by adenoviral vector to overcome the low transfection efficiency of plasmid DNA. The results showed that the expression of SARS-spike protein gene was markedly reduced to 6%. Our studies demonstrate the feasibility of using adenoviral vector to deliver and express the siRNA of specific gene for the therapy of viral infection and other diseases in future.

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