Using Morpholinos to Probe Gene Networks in Sea Urchin.

Using Morpholinos to Probe Gene Networks in Sea Urchin.
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使用吗啡啉探测海胆的基因网络。

DOI:
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发表时间:
2017
影响因子:
--
通讯作者:
Stefan C. Materna
Stefan C. Materna
中科院分区:
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文献类型:
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作者:
Stefan C. Materna

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发育进程的控制过程可以用基因调控网络(GRN)图谱来概括。他们组装的一个关键步骤是对网络候选人的系统干扰。在海胆中,以基因特异性方式干扰表达的最重要方法是应用吗啉代反义寡核苷酸(MO)。 MO 通过与其转录物中的序列互补物结合而发挥作用,导致翻译受阻或剪接发生变化,从而导致功能丧失。尽管这项技术取得了巨大成功,但最近与基因组编辑产生的突变体的比较引起了新的批评,并对其可靠性提出了挑战。与所有基于序列识别的方法一样,MO 很容易发生脱靶结合,这可能导致错误地将表型归因于预期靶标的丢失。然而,海胆发育的缓慢进程使得对胚胎中基因活性的极其详细的研究成为可能。这些丰富的知识与海胆胚胎的简单性相结合,使得能够通过各种不依赖于终端表型的方法对 MO 表型进行仔细分析。本文总结了 MO 在探测 GRN 中的使用以及确保其特异性应采取的步骤。
The control processes that underlie the progression of development can be summarized in maps of gene regulatory networks (GRNs). A critical step in their assembly is the systematic perturbation of network candidates. In sea urchins the most important method for interfering with expression in a gene-specific way is application of morpholino antisense oligonucleotides (MOs). MOs act by binding to their sequence complement in transcripts resulting in a block in translation or a change in splicing and thus result in a loss of function. Despite the tremendous success of this technology, recent comparisons to mutants generated by genome editing have led to renewed criticism and challenged its reliability. As with all methods based on sequence recognition, MOs are prone to off-target binding that may result in phenotypes that are erroneously ascribed to the loss of the intended target. However, the slow progression of development in sea urchins has enabled extremely detailed studies of gene activity in the embryo. This wealth of knowledge paired with the simplicity of the sea urchin embryo enables careful analysis of MO phenotypes through a variety of methods that do not rely on terminal phenotypes. This article summarizes the use of MOs in probing GRNs and the steps that should be taken to assure their specificity.
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