Pea early-browning virus-mediated genome editing via the CRISPR/Cas9 system in Nicotiana benthamiana and Arabidopsis
Pea early-browning virus-mediated genome editing via the CRISPR/Cas9 system in Nicotiana benthamiana and Arabidopsis
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DOI：
10.1016/j.virusres.2017.10.009
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发表时间：
2018-01-15
期刊：
影响因子：
5
通讯作者：
Mahfouz, Magdy M.
中科院分区：
文献类型：
--
作者：
Ali, Zahir;Eid, Ayman;Mahfouz, Magdy M.
聚类规则间隔回文重复序列(CRISPR)/CRISPR相关（Cas9）系统实现了多种植物物种的高效基因组工程。然而，将基因组工程试剂（如单导RNA (sgRNA)）递送到植物细胞中仍然具有挑战性。在这里，我们报道了烟草响铃病毒（TRV）和豌豆早褐变病毒（PEBV）的工程，将一个或多个sgrna传递到过表达含有Cas9的核定位信号的benthamiana和Arabidopsis thaliana （col0）植物中。我们的数据表明，TRV和PEBV可以将sgrna传递到接种和系统叶片中，从而导致目标基因组位点的突变。此外，在N. benthamiana中，基于pebv的sgRNA递送比基于trv的递送导致更多的靶向突变。我们的数据表明，TRV和PEBV可以促进植物基因组工程，并可用于在不同植物物种中产生功能分析和其他生物技术应用的靶向突变。关键信息：基因组工程试剂进入植物细胞具有挑战性和低效率，这限制了该技术在许多植物物种中的应用。TRV和PEBV等RNA病毒为系统递送sgrna进行靶向基因组修饰提供了有效的工具。
The clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated (Cas9) system has enabled efficient genome engineering in diverse plant species. However, delivery of genome engineering reagents, such as the single guide RNA (sgRNA), into plant cells remains challenging. Here, we report the engineering of Tobacco rattle virus (TRV) and Pea early browning virus (PEBV) to deliver one or multiple sgRNAs into Nicotiana benthamiana and Arabidopsis thaliana (Col-0) plants that overexpress a nuclear localization signal containing Cas9. Our data showed that TRV and PEBV can deliver sgRNAs into inoculated and systemic leaves, and this resulted in mutagenesis of the targeted genomic loci. Moreover, in N. benthamiana, PEBV-based sgRNA delivery resulted in more targeted mutations than TRV-based delivery. Our data indicate that TRV and PEBV can facilitate plant genome engineering and can be used to produce targeted mutations for functional analysis and other biotechnological applications across diverse plant species.Key message: Delivery of genome engineering reagents into plant cells is challenging and inefficient and this limit the applications of this technology in many plant species. RNA viruses such as TRV and PEBV provide an efficient tool to systemically deliver sgRNAs for targeted genome modification.