A virus-based miRNA suppression (VbMS) system for miRNA loss-of-function analysis in plants.
A virus-based miRNA suppression (VbMS) system for miRNA loss-of-function analysis in plants.
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DOI：
10.1002/biot.201300523
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发表时间：
2014-05
影响因子：
4.7
通讯作者：
Fei Yan;Weixia Guo;Gentu Wu;Yuwen Lu;Jiejun Peng;Hongying Zheng;Lin Lin-Lin;Jianping Chen
中科院分区：
文献类型：
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作者：
Fei Yan;Weixia Guo;Gentu Wu;Yuwen Lu;Jiejun Peng;Hongying Zheng;Lin Lin-Lin;Jianping Chen
microRNA（miRNAs）在植物发育和防御病原体中起着关键作用。为了确定单个miRNA的功能，通常通过在转基因植物中过表达特定的miRNA来进行功能获得性分析，并且已经证明非常有效。通过靶模拟方法的功能丧失分析现在也越来越多地被使用。在转基因植物中表达的模拟物隔离特定的miRNA并导致阐明miRNA功能的改变的表型。然而，获得转基因植物需要时间。为了避免使用转基因植物，我们已经开发了一种基于烟草脆裂病毒载体的基于病毒的miRNA抑制系统（VbMS）。将miR156、miR319或miR164的靶模拟序列引入病毒基因组RNA中，然后将其接种到拟南芥植物中。所得表型与先前来自转基因植物的报道一致，并且miRNA的靶标的表达也增加，表明miRNA的活性已被抑制。本文开发的VbMS被验证用于植物中miRNA的功能丧失分析。此外，由于只需要简单的农杆菌渗透而不是转化，VbMS适合于大规模的方法在植物中的miRNA功能分析。
MicroRNAs (miRNAs) play key roles in plant development and defense against pathogens. To establish the function of individual miRNAs, gain-of-function analysis is usually done by overexpressing a specific miRNA in transgenic plants and has proved very effective. Loss-of-function analysis by the target mimic method is now also increasingly being used. The mimics expressed in the transgenic plants sequester a specific miRNA and lead to changed phenotypes that elucidate miRNA function. However, it takes time to obtain the transgenic plants. To avoid using transgenic plants, we have developed a virus-based miRNA suppression system (VbMS) based on a Tobacco rattle virus vector. The target mimic sequences of miR156, miR319, or miR164 were introduced into the viral genomic RNA, which was then inoculated to Arabidopsis thaliana plants. The resulting phenotypes were consistent with previous reports from transgenic plants, and the expression of targets of the miRNAs was also increased showing that the activity of the miRNAs had been inhibited. VbMS developed here is validated for loss-of-function analysis of miRNA in plants. Moreover, since only simple agroinfiltration rather than transformation is needed, VbMS is suitable for large-scale approaches to miRNA function analysis in plants.