PDF(Pubmed)
Abstract：
Antisense oligodeoxynucleotides (ASOs) have long been used to selectively inhibit or modulate gene expression at the RNA level, and some ASOs are approved for clinical use. However, the practicability of antisense technologies remains limited by the difficulty of reliably predicting the sites accessible to ASOs in complex folded RNAs. Recently, we applied a plant-based method that reproduces RNA-induced RNA silencing in vitro to reliably identify sites in target RNAs that are accessible to small interfering RNA (siRNA)-guided Argonaute endonucleases. Here, we show that this method is also suitable for identifying ASOs that are effective in DNA-induced RNA silencing by RNases H. We show that ASOs identified in this way that target a viral genome are comparably effective in protecting plants from infection as siRNAs with the corresponding sequence. The antiviral activity of the ASOs could be further enhanced by chemical modification. This led to two important conclusions: siRNAs and ASOs that can effectively knock down complex RNA molecules can be identified using the same approach, and ASOs optimized in this way could find application in crop protection. The technology developed here could be useful not only for effective RNA silencing in plants but also in other organisms.
摘要：
反义寡脱氧核苷酸（ASO）长期以来被用于在RNA水平上选择性地抑制或调节基因表达。一些ASO被批准用于临床。然而,反义技术的实用性仍然受到难以可靠预测复杂折叠RNA中ASO可接近的位点的限制。最近,我们应用了一种基于植物的方法,该方法在体外复制RNA诱导的RNA沉默,以可靠地鉴定靶RNA中小干扰RNA(siRNA)指导的Argonaute内切核酸酶可接近的位点.这里,我们表明，该方法也适用于鉴定在RNA酶H的DNA诱导的RNA沉默中有效的ASO。我们表明，以这种方式鉴定的靶向病毒基因组的ASO在保护植物免受感染方面相对有效，作为具有相应序列的siRNA。通过化学修饰可以进一步增强ASO的抗病毒活性。这导致了两个重要的结论：可以使用相同的方法鉴定可以有效敲除复杂RNA分子的siRNA和ASO，以这种方式优化的ASO可以在作物保护中找到应用。这里开发的技术不仅可用于植物中的有效RNA沉默，也可用于其他生物体。
