A resource for large-scale RNA-interference-based screens in mammals
A resource for large-scale RNA-interference-based screens in mammals
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DOI：
10.1038/nature02370
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发表时间：
2004-03-25
期刊：
影响因子：
64.8
通讯作者：
Hannon, GJ
中科院分区：
文献类型：
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作者：
Paddison, PJ;Silva, JM;Hannon, GJ
在哺乳动物细胞中使用小干扰RNA（siRNA）和短发夹RNA（shRNA）通过RNA干扰（RNAi）进行基因沉默已成为有价值的遗传工具（1-10）。在此，我们报告了针对9，610个人类和5，563个小鼠基因的shRNA表达文库的构建和应用。该文库目前由包含在多功能载体内的约28，000个序列验证的shRNA表达盒组成，其允许shRNA盒包装在逆转录病毒中，通过DNA“条形码”在混合细胞群体中追踪，并通过细菌交配穿梭至定制载体。为了验证文库，我们使用了一个基因筛选，旨在报告人类蛋白酶体功能的缺陷。我们的研究结果表明，我们的大规模RNAi文库可以用于特定的，在哺乳动物的遗传应用，并将成为一个有价值的资源，基因分析和发现。
Gene silencing by RNA interference (RNAi) in mammalian cells using small interfering RNAs (siRNAs) and short hairpin RNAs (shRNAs) has become a valuable genetic tool(1-10). Here, we report the construction and application of a shRNA expression library targeting 9,610 human and 5,563 mouse genes. This library is presently composed of about 28,000 sequence-verified shRNA expression cassettes contained within multi-functional vectors, which permit shRNA cassettes to be packaged in retroviruses, tracked in mixed cell populations by means of DNA 'bar codes', and shuttled to customized vectors by bacterial mating. In order to validate the library, we used a genetic screen designed to report defects in human proteasome function. Our results suggest that our large-scale RNAi library can be used in specific, genetic applications in mammals, and will become a valuable resource for gene analysis and discovery.