Short hairpin RNAs (shRNAs) induce sequence-specific silencing in mammalian cells
Short hairpin RNAs (shRNAs) induce sequence-specific silencing in mammalian cells
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DOI：
10.1101/gad.981002
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发表时间：
2002-04-15
影响因子：
10.5
通讯作者：
Conklin, DS
中科院分区：
文献类型：
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作者：
Paddison, PJ;Caudy, AA;Conklin, DS
RNA干扰(RNAi)首先在秀丽线虫中被发现，是对外源双链RNA(DsRNA)的生物反应，导致序列特异性基因沉默。RNAi是一种保守的调控基序，广泛存在于真核生物中。最近，我们和其他人已经证明，内源编码的基因沉默触发器通过RNAi机制的元件来调节蛋白质编码基因的表达。这些小的临时RNA(StRNAs)被转录为短发夹状前体(类似于70nT)，被DICER加工成活性的21nT的RNAs，并通过碱基配对相互作用识别目标mRNAs。在这里，我们展示了短发夹状RNA(ShRNAs)可以被改造成在培养的果蝇和哺乳动物细胞中抑制所需基因的表达。ShRNAs可以从外源合成，也可以在体内从RNA聚合酶III启动子转录，从而允许构建连续的细胞系或转基因动物，在其中RNAi实施稳定的和可遗传的基因沉默。
RNA interference (RNAi) was first recognized in Caenorhabditis elegans as a biological response to exogenous double-stranded RNA (dsRNA), which induces sequence-specific gene silencing. RNAi represents a conserved regulatory motif, which is present in a wide range of eukaryotic organisms. Recently, we and others have shown that endogenously encoded triggers of gene silencing act through elements of the RNAi machinery to regulate the expression of protein-coding genes. These small temporal RNAs (stRNAs) are transcribed as short hairpin precursors (similar to70 nt), processed into active, 21-nt RNAs by Dicer, and recognize target mRNAs via base-pairing interactions. Here, we show that short hairpin RNAs (shRNAs) can be engineered to suppress the expression of desired genes in cultured Drosophila and mammalian cells. shRNAs can be synthesized exogenously or can be transcribed from RNA polymerase III promoters in vivo, thus permitting the construction of continuous cell lines or transgenic animals in which RNAi enforces stable and heritable gene silencing.