RNA interference (RNAi) has become an integral part of mainstream research due to its versatility and ease of use. However, the potential nontarget effects associated with double‐stranded RNAs (dsRNA) are poorly understood. To explore this, we used dsRNAs targeting the inhibitor of apoptosis (iap) gene from nine insect species and assayed their possible nontarget effects. For each assay, we used a control (dsRNA targeting the gene coding for green fluorescent protein, GFP) and a species‐specific dsRNA targeting nine iap genes in insect species to evaluate target gene knockdown efficiency, apoptosis phenotype in cells and mortality in insects. Our results revealed that dsIAP efficiently knocks down iap gene expression and induces apoptosis phenotype and mortality in target insect species. In contrast, no significant knockdown of the iap gene expression, apoptosis phenotypes, or mortality were detected in cell lines developed from nontarget insects or nontarget insects treated with dsIAPs. Interestingly, even among closely related insects such as stink bugs, Nezara viridula, Halyomorpha halys, and Murgantia histrionica, with substantial sequence similarity among iap genes from these insects, no significant nontarget effects of dsIAP were observed under the conditions tested. These data demonstrate no significant nontarget effects for dsIAPs and suggest that the threat of nontarget effects of RNAi technology may not be substantial.

中文翻译：

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靶向凋亡基因抑制剂的双链 RNA 没有显示出显着的跨物种活性。

由于其多功能性和易用性，RNA 干扰 (RNAi) 已成为主流研究中不可或缺的一部分。然而，人们对与双链 RNA (dsRNA) 相关的潜在非目标效应知之甚少。为探索这一点，我们使用 dsRNAs 靶向来自九种昆虫的细胞凋亡抑制剂 ( iap ) 基因，并分析了它们可能的非靶点效应。对于每个测定，我们使用对照（靶向绿色荧光蛋白 GFP 基因编码的 dsRNA）和靶向昆虫物种中九个iap基因的物种特异性 dsRNA 来评估靶基因敲低效率、细胞凋亡表型和昆虫死亡率。我们的结果表明 dsIAP 有效地击倒了iap基因表达并诱导目标昆虫物种的细胞凋亡表型和死亡率。相反，在从非目标昆虫或用 dsIAP 处理的非目标昆虫开发的细胞系中，未检测到iap基因表达、细胞凋亡表型或死亡率的显着降低。有趣的是，即使在臭虫、Nezara viridula、Halyomorpha halys和Murgantia histrionica等密切相关的昆虫中， iap之间也具有显着的序列相似性来自这些昆虫的基因，在测试条件下没有观察到 dsIAP 的显着非目标效应。这些数据表明 dsIAP 没有显着的非目标效应，并表明 RNAi 技术的非目标效应威胁可能并不大。