Genome editing in maize directed by CRISPR-Cas9 ribonucleoprotein complexes.
Genome editing in maize directed by CRISPR-Cas9 ribonucleoprotein complexes.
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DOI：
10.1038/ncomms13274
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发表时间：
2016-11-16
影响因子：
16.6
通讯作者：
Cigan, A. Mark
中科院分区：
文献类型：
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作者：
Svitashev, Sergei;Schwartz, Christine;Lenderts, Brian;Young, Joshua K.;Cigan, A. Mark
靶向DNA双链断裂已被证明可以显着提高基因组编辑的频率和精度。在过去的二十年中，已经开发了几种双链断裂技术。CRISPR-Cas9因其简单、高效和多功能性而迅速成为基因组编辑的首选技术。目前，植物中的基因组编辑主要依赖于以DNA载体的形式递送双链断裂试剂。在这里，我们报告了将预组装的Cas9-gRNA核糖核蛋白生物射弹递送到玉米胚细胞中以及具有突变和编辑的等位基因的植物的再生。使用这种方法的交付，我们还证明了DNA和选择标记的基因突变，在玉米和恢复植物突变等位基因在高频率。这些结果为加速各种作物品种的育种实践提供了新的机会。 植物中的基因组编辑通常需要从稳定转化的质粒DNA表达Cas9和指导RNA。在这里，作者表明，通过生物射弹将Cas9-指导RNA复合物作为核糖核蛋白递送到玉米胚后，可以实现成功的编辑。
Targeted DNA double-strand breaks have been shown to significantly increase the frequency and precision of genome editing. In the past two decades, several double-strand break technologies have been developed. CRISPR–Cas9 has quickly become the technology of choice for genome editing due to its simplicity, efficiency and versatility. Currently, genome editing in plants primarily relies on delivering double-strand break reagents in the form of DNA vectors. Here we report biolistic delivery of pre-assembled Cas9–gRNA ribonucleoproteins into maize embryo cells and regeneration of plants with both mutated and edited alleles. Using this method of delivery, we also demonstrate DNA- and selectable marker-free gene mutagenesis in maize and recovery of plants with mutated alleles at high frequencies. These results open new opportunities to accelerate breeding practices in a wide variety of crop species. Genome editing in plants typically requires the expression of Cas9 and guide RNA from stably transformed plasmid DNA. Here, the authors show that successful editing can be achieved after delivery of the Cas9-guide RNA complex as a ribonucleoprotein to maize embryos via biolistics.
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