DNA interference:: a simple and efficient gene-silencing system for high-throughput functional analysis in the fern Adiantum
DNA interference:: a simple and efficient gene-silencing system for high-throughput functional analysis in the fern Adiantum
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DOI：
10.1093/pcp/pch186
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发表时间：
2004-11-01
影响因子：
4.9
通讯作者：
Wada, M
中科院分区：
文献类型：
--
作者：
Kawai-Toyooka, H;Kuramoto, C;Wada, M
RNA干扰（RNAi）已成为确定基因功能的有力工具，并在各种生物中使用。由于需要产生双链RNA（dsRNA）作为RNAi的诱导物，因此RNAi诱导构建体的制备有些繁琐和耗时，特别是对于全基因组分析中使用的数千个基因。为了克服这些问题，我们已经开发了一种更方便的基因沉默方法在铁线蕨使用双链DNA（dsDNA）作为模式系统的功能分析在植物中。将与内源基因同源的dsDNA片段递送到配子体细胞中可以诱导序列特异性基因沉默。由于它只需要与靶基因同源的dsDNA片段，PCR扩增的片段足以触发基因沉默。对于转化的植物，已经实现了>90%的最大基因沉默效率。此外，同时转移对应于多个基因的dsDNA片段对单个基因仍具有沉默效应。我们称这种方法为“DNA干扰”。
RNA interference (RNAi) has become a powerful tool for determining gene function and is used in a wide variety of organisms. Since it is necessary to generate double-stranded RNA (dsRNA) as an inducer for RNAi, preparation of RNAi-inducing constructs is somewhat cumbersome and time consuming, especially for the thousands of genes used in a genome-wide analysis. To overcome these problems, we have developed a more convenient gene-silencing method in the fern Adiantum using double-stranded DNA (dsDNA) as a model system for functional analysis in plants. Delivery of dsDNA fragments homologous to an endogenous gene into gametophytic cells can induce sequence-specific gene silencing. As it only requires dsDNA fragments homologous to a target gene, PCR-amplified fragments are enough to trigger gene silencing. Maximum gene silencing efficiencies of >90% have been achieved for transformed plants. In addition, simultaneous transfer of dsDNA fragments corresponding to multiple genes still has a silencing effect for individual genes. We term this approach 'DNA interference'.