Turning Up the Temperature on CRISPR: Increased Temperature Can Improve the Editing Efficiency of Wheat Using CRISPR/Cas9.
Turning Up the Temperature on CRISPR: Increased Temperature Can Improve the Editing Efficiency of Wheat Using CRISPR/Cas9.
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DOI：
10.3389/fpls.2020.583374
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发表时间：
2020
影响因子：
5.6
通讯作者：
Wallington EJ
中科院分区：
文献类型：
--
作者：
Milner MJ;Craze M;Hope MS;Wallington EJ
CRISPR/Cas9技术的应用已经改变了我们靶向和编辑基因组指定区域的能力。它对任何生物体的广泛适应性使我们对许多生物过程的理解取得了无数进步。目前的许多工具都是为简单的植物系统（如二倍体物种）设计的，然而，在作物物种中有效部署需要更高的编辑效率，因为这些物种通常包含多倍体基因组。在这里，我们研究了温度的作用，以了解是否可以在小麦中提高CRISPR/Cas9编辑效率。最近的研究发现，在较高温度下的植物生长可能会增加突变率，他们用小麦中两种不同启动子表达的Cas9进行了测试。当Cas9酶由ZmUbi启动子驱动而非OsActin驱动时，提高组织培养温度或种子萌发和生长早期的温度会增加小麦的突变频率。相比之下，由OsActin启动子驱动的Cas9表达并没有增加转化系或转化过程中检测到的突变。这些结果表明，在多倍体谷物物种中，通过简单地改变生长条件来促进增加突变以产生纯合或零敲除，CRISPR/Cas9编辑效率可以显着提高。
The application of CRISPR/Cas9 technologies has transformed our ability to target and edit designated regions of a genome. It’s broad adaptability to any organism has led to countless advancements in our understanding of many biological processes. Many current tools are designed for simple plant systems such as diploid species, however, efficient deployment in crop species requires a greater efficiency of editing as these often contain polyploid genomes. Here, we examined the role of temperature to understand if CRISPR/Cas9 editing efficiency can be improved in wheat. The recent finding that plant growth under higher temperatures could increase mutation rates was tested with Cas9 expressed from two different promoters in wheat. Increasing the temperature of the tissue culture or of the seed germination and early growth phase increases the frequency of mutation in wheat when the Cas9 enzyme is driven by the ZmUbi promoter but not OsActin. In contrast, Cas9 expression driven by the OsActin promoter did not increase the mutations detected in either transformed lines or during the transformation process itself. These results demonstrate that CRISPR/Cas9 editing efficiency can be significantly increased in a polyploid cereal species with a simple change in growth conditions to facilitate increased mutations for the creation of homozygous or null knock-outs.
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