Genorne-wide RNAi screening in Caenorhabditis elegans
Genorne-wide RNAi screening in Caenorhabditis elegans
复制标题
DOI：
10.1016/s1046-2023(03)00050-1
复制
发表时间：
2003-08-01
期刊：
影响因子：
4.8
通讯作者：
Ahringer, J
中科院分区：
文献类型：
--
作者：
Kamath, RS;Ahringer, J
In Caenorhabditis elegans, introduction of double-stranded RNA (dsRNA) results in the specific inactivation of an endogenous gene with corresponding sequence;这种技术称为 RNA 干扰 (RNAi)。先前已经表明，RNAi 可以通过将 dsRNA 直接显微注射到成年雌雄同体蠕虫中、将蠕虫浸泡在 dsRNA 溶液中或通过喂养表达靶基因 dsRNA 的大肠杆菌来进行。我们开发了一种简单的优化方案，利用第三种 dsRNA 引入模式，即喂食 RNAi，可以快速有效地分析线虫中的基因功能。此外，我们构建了一个细菌菌株文库，对应于线虫中估计 19,000 个预测基因中的大约 86%，并用它来进行基因功能的全基因组分析。该库是公开可用的、可重复使用的资源，允许快速大规模 RNAi 实验。我们使用该库对秀丽隐杆线虫的基因功能进行全基因组分析。在这里，我们描述了用于细菌文库构建和使用 RNAi 喂养进行线虫高通量筛选的方案。 (C) 2003 年爱思唯尔科学（美国）。版权所有。
In Caenorhabditis elegans, introduction of double-stranded RNA (dsRNA) results in the specific inactivation of an endogenous gene with corresponding sequence; this technique is known as RNA interference (RNAi). It has previously been shown that RNAi can be performed by direct microinjection of dsRNA into adult hermaphrodite worms, by soaking worms in a solution of dsRNA, or by feeding worms Escherichia coli expressing target-gene dsRNA. We have developed a simple optimized protocol exploiting this third mode of dsRNA introduction, RNAi by feeding, which allows rapid and effective analysis of gene function in C elegans. Furthermore, we have constructed a library of bacterial strains corresponding to roughly 86% of the estimated 19,000 predicted genes in C elegans, and we have used it to perform genome-wide analyses of gene function. This library is publicly available, reusable resource allowing for rapid large-scale RNAi experiments. We have used this library to perform genome-wide analyses of gene function in C elegans. Here, we describe the protocols used for bacterial library construction and for high-throughput screening in C elegans using RNAi by feeding. (C) 2003 Elsevier Science (USA). All rights reserved.