对于靶向基因沉默，RNA干扰（RNAi)具有易于操作、快速结果、高效率、广泛适用于各种细胞类型的多重优势。其具有瞬转效应和剂量依赖效应，这点与小分子非常相似。然而，目前的siRNA试剂的特异性和基因沉默效率不是很稳定，阻碍了它们作为药物发现和基因研究工具的应用。

 

可靠表型的特异性基因沉默工具

siRNA pools（siPOOLs）是一款经过优化设计的、高复杂性的、包含30条siRNA的混合物，经证明可有效消除脱靶效应，提高了结果的可靠性（Hannus et al., 2014）。 Pack Hunter (pooling) 方法通过将当个siRNA的浓度稀释到刺激表型的阈值以下来对抗单个siRNA的脱靶。 借助专有的设计算法，siPOOLs中的siRNA序列经过优化，以实现最大的转录本覆盖率，高效杂交并对旁系同源基因进行过滤，从而实现高效和特异性的基因沉默。
 

siPOOLs产品优势

• 使用简单快捷：siPOOLs与多种转染试剂兼容，几天内就能看到结果。
• 高度特异性且有效：siPOOLs在标准细胞系中将脱靶率降低5-25 倍，且在1-3 nM下实现基因敲低率≥70%。
• 一致的表型：由序列非依赖性的siPOOL产生的表型高度一致。
• 确保经过检验：通过RT-qPCR进行siPOOL验证，如果在最佳转染下敲低率低于70%，则有可能重新设计。
• 使用专业数据库的注释进行定制设计：专业的设计，确保优化热力学特性且避免旁系同源基因。
• HPLC纯化且无毒：所有siPOOL均经过HPLC纯化，可降低污染物和副作用的风险。

关键问题：siRNAs的脱靶效应

siRNA通常与靶RNA转录本互补结合，通过RNAi机制指导其降解。脱靶效应主要是由siRNA模拟内源性基因调节因子microRNA（miRNA）引起的。由于miRNA只需要6个碱基种子匹配到3'非翻译区（UTR），即可触发转录本下调。因此，siRNA在通过这种机制发挥作用时，可能会改变许多意想不到的靶标基因的表达。
 

siPOOLs如何提高特异性？

单个siRNA或含有3-4个siRNA的低复杂度siRNA库，常常击中多个脱靶基因，并表现出易变的靶基因敲低效率。 siPOOL是高度复杂且确定的30个siRNA池，每个siRNA以皮摩尔工作浓度存在。因此，siPOOLs ：
1. 有效稀释了每个siRNA的脱靶特征，提高了靶向特异性。
2. 确保了靶基因的协同敲低，产生更稳健、更可靠的结果。

 

使用siPOOL具有更高的特异性

特异性好的试剂，应仅影响其靶标基因。

HeLa细胞的微阵列表达谱分析显示，单个siRNA可以诱导许多脱靶基因（红点），而针对同一靶基因（绿点）的siPOOL，或许也包含有非特异性的siRNA，但是大大降低了脱靶效应。
 

通过siPOOL实现更好的重现性和有效的敲低效率

普通siRNA的敲低效率差异很大。

与单个siRNA相比，针对同一基因的siPOOLs具有相似的敲低效率，表明具有更高的稳健性和可重复性（左图和中图：靶RNA水平的实时定量PCR测量的相关图）。 siPOOLs也表现出有效的基因敲低。在常用的细胞系中，许多基因通常在1 nM浓度下实现75-98%的基因敲低效率（右图）。
 

您可以信赖的表型

如果RNAi效果可靠且特异，靶向同一基因的两种试剂应产生相似的表型。

每个基因分别使用两个siPOOL池，一个来自我们的人类激酶siPOOL文库，一个是市售的每个基因含3条siRNAs的文库，科学家在A549细胞中筛选了36个基因并检测了细胞活力，结果表明siPOOLs产生了较好的一致性表型。

siTOOLs Biotech由Michael Hannus博士和Gunter Meister教授于2013年9月成立，由德国雷根斯堡大学 和Intana Bioscience制药服务公司联合运营。目前推出有3个产品系列：（1）siPOOLs：一种siRNA混合物，有效增加基因沉默特异性，“稀释”传统RNA干扰试剂的脱靶效应。（2）raPOOLs：一种稳健的RNA亲和纯化方案，适用于基因功能和相互作用的生物化学研究。（3）riboPOOLs：适合任意物种的核糖体RNA去除试剂，高效且经济。

siTOOLs Biotech始终致力于帮助学术研究人员、科学家、制药/生物技术公司和RNAi筛选人员。目前，已帮助许多论文发表在《自然》、《细胞》、《自然医学》、《科学报告》等顶级学术期刊上。期待与您的合作，我们将为您的RNA研究提供量身定制的分子工具。

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