RNA干涉的研究进展

生物体内导入双链RNA后会引起体内同源基因特异性的沉默,这种现象称为RNA干涉,本主要介绍RNA干涉的研究历史,作用机制和应用等方面的情况。     

RNA干涉分子机制研究进展

RNA干涉（RNA interference,RNAi）是生物体内的一种通过双链RNA(dsRNA)来抵抗病毒入侵和抑制转座子活动的自然机制.双链RNA与同源mRNA互补结合而使特定基因失活,这一过程已经在包括拟南芥、线虫和真菌等多种模式生物中得到揭示.近来研究表明,21～25 nt的小干涉RNA（small interference RNA, siRNA）可介导哺乳动物细胞特异性基因沉默.RNAi具有高效性和高度特异性,可能成为关闭基因的新技术而在基因功能研究和疾病基因治疗中发挥重要作用.     

RNA干涉与干细胞

RNA干涉(RNAi)现象普遍存在于生物体细胞中,在理论上已清楚其分子机制,为干细胞研究提供了新的方法。现从RNAi的分子机制、干细胞中的RNAi现象、研究干细胞RNAi效应的方法以及小分子干涉RNA(siRNA)干涉干细胞特异功能基因的检测方法等方面进行了综述。表明应用RNAi技术研究基因功能和干细胞维持及定向分化的调控具有广阔的发展前景。     

RNA干涉及其应用前景

RNA干涉是指由特定双链RNA(dsRNA)引起的转录后基因沉默现象。研究表明,Dicer断裂dsRNA产生的小干涉RNA可以抑制哺乳动物体细胞和胚胎中的基因的表达。RdRP在扩增RNAi中起着关键性的作用,RdRP活性复制较长的触发性dsRNA或以一种非引物的方式复制短的siRNA,即以siRNA为引物的RdRP反应使靶mRNA转变为dsRNA,同时复制触发性dsRNA。所有的产物又可作为Dicer的底物,起始RdRP级联反应。本文综述了RNAi可能的作用机制,并对RNAi在分析功能基因组、药物治疗等方面的应用前景进行了展望。     

RNA干涉与基因沉默

双链RNA介导的遗传干涉的机制是1998年发现的,它通过双链RNA的介导特异性地降解相应序列的mRNA,从而导致转录后水平的基因沉默,到目前为止的真菌,拟南芥,线虫,锥虫,水螅,涡虫,果蝇,斑马鱼,小鼠等真核生物中都发现存在这一基因沉默机制,目前的研究表明,RNA干涉与植物中的共抑制（Cosuppression),真菌中的基因压制（quelling)很可能具有共同的基本分子机制,这也说明,很可能在进化的很早期阶段,生物就获得了这种机制,RNA干涉对于抵抗病毒入侵,抑制转座子活动等具有重要作用,对于生物体的发育和基因调控可能也有重要作用。     

RNA干涉的最新研究进展

RNA干涉 (RNAi)是将双链RNA(dsRNA)导入细胞引起特异基因mRNA降解的一种细胞反应过程 .它是转录后基因沉默 (PTGS)的一种 .RNAi在生物界中广泛存在 .RNAi发生过程主要分为 3个阶段 :起始阶段 ,扩增阶段 ,效应阶段 .RNAi在维持基因组稳定、保护基因组免受外源核酸侵入、基因表达调控等方面发挥重要生物学作用 .RNAi作为基因沉默的一个工具 ,已被广泛用于基因功能研究、基因治疗和新药研究与开发等方面 .     

RNA干涉技术(RNAi)应用研究进展

RNA干涉是通过双链RNA的介导特异性地降解相应序列的mRNA,从而导致转录后水平的基因沉默,这一过程在拟南芥、线虫和真菌等多种模式植物中发现。RNAi是研究多种生物基因功能的有效手段,本文综述了RNA干涉的分子机理及其应用研究进展。     

RNA干涉技术

RNA干涉(RNAi)技术是利用一些小的双链RNA来高效、特异地阻断体内特定基因的表达,并促使mRNA降解,从而诱使细胞表现出特定基因缺失的表型。本从RNAi技术的历史、作用机制、研究策略、研究现状及应用前景等几个方面进行了综述,预测RNAi将会给基因治疗的发展带来新的希望。     

通过RNA干涉沉默SARS病毒的研究前景

RNA干涉(RNAi)是双链RNA介导的转录后基因沉默作用的重要机制之一。RNAi在后基因组时代的基因功能研究和药物开发中具有广阔应用前景。本综述了RNAi在抗SARS病毒药物研究中的应用前景。     

Dynamics of gene silencing by RNA interference

The large number of candidate genes identified by modern high-throughput technologies require efficient methods for generating knockout phenotypes or gene silencing in order to study gene function. RNA interference (RNAi) is an efficient method that can be used for this purpose. Effective gene silencing by RNAi depends on a number of important parameters, including the dynamics of gene expression and the RNA dose. Using mouse hepatoma cells, we detail some of the principal characteristics of RNAi as a tool for gene silencing, such as the RNA dose level, RNA complex exposure time, and the time of transfection relative to gene induction, in the context of silencing a green fluorescent protein reporter gene. Our experiments demonstrate that different levels of silencing can be attained by modulating the dose level of RNA and the time of transfection and illustrate the importance of a dynamic analysis in designing robust silencing protocols. By quantifying the kinetics of RNAi-based gene silencing, we present a model that may be used to help determine key parameters in more complex silencing experiments and explore alternative gene silencing protocols.     

RNA interference: the story of gene silencing in plants and humans

RNA interference is an exciting field of functional genomics that can silence viral genes. This property of interfering RNA can be used to combat viral diseases of plants as well as animals and humans. It is a short sequence of nucleic acid that can bind to the mRNA of the gene and interferes the process of its expression. It is diverse in occurrence as well as in applications. It occurs from nematodes to fungi and can cause gene silencing in plants, animals and human beings. Small interfering RNAs are used to silence plant viral genes and in production of therapeutic drugs against Hepatitis or Immuno-deficiency viruses in human. In this review, we will discuss the history, mechanism and applications of RNA interference in plant, animal and human research.     

RNA干扰作用研究进展

RNA干扰 (RNAinterference ,RNAi)是指与内源性mRNA编码区某段序列同源的双链RNA分子 (double strandedRNA ,dsRNA)导入细胞时 ,该mRNA发生特异性的降解 ,导致基因表达的沉默。本文介绍RNAi作用的发现、机制和目前使用的产生RNAi的方法     

Moment analysis for kinetics of gene silencing by RNA interference

RNA interference (RNAi) was quantitatively evaluated from a kinetic viewpoint. A simple kinetic evaluation based on moment analysis was proposed, assuming suppression and recovery phases of gene expression. We defined the area under the curve of the inhibitory effect (AUC(IE)) as an index of the total intensity of RNAi and the mean response time of the inhibitory effect (MRT(IE)) as an index of its duration. The proposed kinetic analysis helps to understand the RNAi effect in a quantitative and time-dependent manner, which will be beneficial for designing RNAi-based gene silencing for both experimental and therapeutic purposes.     

Myostatin gene silencing by RNA interference in chicken embryo fibroblast cells

Myostatin (MSTN), a member of transforming growth factor-β (TGF-β) superfamily, is a negative regulator of the skeletal muscle growth, and suppresses the proliferation and differentiation of myoblast cells. Dysfunction of MSTN gene either by natural mutation or genetic manipulation (knockout or knockdown) has been reported to interrupt its proper function and to increase the muscle mass in many mammalian species. RNA interference (RNAi) mediated by small interfering RNAs (siRNAs) or short hairpin RNAs (shRNAs) has become a powerful tool for gene knockdown studies. In the present study transient silencing of MSTN gene in chicken embryo fibroblast cells was evaluated using five different shRNA expression constructs. We report here up to 68% silencing of myostatin mRNA using these shRNA constructs in transiently transfected fibroblasts (p<0.05). This was, however, associated with induction of interferon responsive genes (OAS1, IFN-β) (3.7-64 folds; p<0.05). Further work on stable expression of antimyostatin shRNA with minimum interferon induction will be of immense value to increase the muscle mass in the transgenic animals.     

Different effects on ACC oxidase gene silencing triggered by RNA interference in transgenic tomato

RNA interference (RNAi) is a potent trigger for specific gene silencing of expression in a number of organisms and is an efficient way of shutting down gene expression. 1-Aminocyclopropane-1-carboxylate (ACC) oxidase catalyzes the oxidation of ACC to ethylene, a plant growth regulator that plays an important role in the tomato ripening process. In this research, to produce double-stranded (ds)RNA of tomato ACC oxidase, we linked the sense and antisense configurations of DNA fragments with 1,002-bp or 7-nt artificially synthesized fragments, respectively, and then placed these under the control of a modified cauliflower mosaic virus 35S promoter. The dsRNA expression unit was successfully introduced into tomato cultivar Hezuo 906 by Agrobacterium tumefaciens-mediated transformation. Molecular analysis of 183 transgenic plants revealed that the dsRNA unit was integrated into the tomato genome. With respect to the construct with the 1,002-bp linker, the severity of phenotypes indicated that 72.3% of the transformed plants had non-RNA interference, about 18.1% had semi-RNA interference, and only 9.6% had full-RNA interference. However when the construct with the 7-nt linker was used for transformation, the results were 13.0%, 18.0%, and 69.0%, respectively, indicating that the short linker was more efficient in RNAi of transgenic tomato plants. When we applied this fast way of shutting down the ACC oxidase gene, transgenic tomato plants were produced that had fruit which released traces of ethylene and had a prolonged shelf life of more than 120 days. The RNA and protein analyses indicated that there was non-RNA interference, semi-RNA interference and full-RNA interference of ACC oxidase in the transgenic tomato plants.     

Autocidal control of ticks by silencing of a single gene by RNA interference

Ticks impact human and animal health worldwide and new control methods are needed to circumvent drawbacks of tick control by acaricide application including selection of drug resistant ticks and environmental pollution. Using RNA interference we silenced the expression of a single gene, subolesin, and produced ticks with diminished reproductive performance and prevented successful mating and production of viable offspring. We propose a sterile acarine technique (SAT) for reduction of tick populations by release of subolesin-silenced ticks. Conservation of subolesin among tick species suggests that SAT may be useful for control of many medically and economically important tick species.     

Tissue-specific gene silencing by RNA interference in the whitefly Bemisia tabaci (Gennadius)

The hemipteran whitefly Bemisia tabaci (Gennadius) species complex and the plant viruses they transmit pose major constraints to vegetable and fiber production, worldwide. The whitefly tissue- and developmental-specific gene expression has not been exhaustively studied despite its economic importance. In 2002, a functional genomic project was initiated, which generated several thousands expressed sequence tags (ESTs) and their sequence. This project provides the basic information to design experiments aimed at understanding and manipulating whitefly gene expression. In this communication, for the first time we provide evidence that the RNA interference mechanism discovered in many organisms, including in Hemiptera, is active in B. tabaci. By injecting into the body cavity long double-stranded RNA (dsRNA) molecules, specifically directed against genes uniquely expressed in the midgut and salivary glands, we were able to significantly inhibit the expression of the targeted mRNA in the different organs. Gene expression levels in RNAi-silenced whiteflies were reduced up to 70% compared to whiteflies injected with buffer or with a green fluorescent protein (GFP)-specific dsRNA. Phenotypic effects were observed in B. tabaci ovaries following dsRNA targeting the whitefly Drosophila chickadee homologue. Disruption of whitefly gene expression opens the door to new strategies aimed at curbing down the deleterious effects of this insect pest to agriculture.