Highly Efficient, Rapid and Co-CRISPR-Independent Genome Editing in Caenorhabditis elegans
Highly Efficient, Rapid and Co-CRISPR-Independent Genome Editing in Caenorhabditis elegans
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DOI：
10.1534/g3.117.300216
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发表时间：
2017-11-01
影响因子：
2.6
通讯作者：
Beg, Asim A.
中科院分区：
文献类型：
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作者：
Prior, Harriet;Jawad, Ali K.;Beg, Asim A.
我们描述了一种使用直接注射CRISPR-Cas9核糖核蛋白在秀丽隐杆线虫中产生点突变的快速且高效的方法。这种多功能方法不需要敏化遗传背景或基于co-CRISPR选择的方法，并且代表了可用于创建基因组点突变的单一策略，无论位置如何。作为原则的证明，我们表明，敲入突变体更忠实地报告变异相关的表型相比，转基因过表达。9个敲入突变体跨越5个基因的数据，表现出高编辑效率（60%），减少筛选工作量（24 F1后代），和快速的时间尺度（4-5天）。这种优化的方法简化了基因组工程，并很容易适应其他模型系统。
We describe a rapid and highly efficient method to generate point mutations in Caenorhabditis elegans using direct injection of CRISPR-Cas9 ribonucleoproteins. This versatile method does not require sensitized genetic backgrounds or co-CRISPR selection-based methods, and represents a single strategy that can be used for creating genomic point mutations, regardless of location. As proof of principle, we show that knock-in mutants more faithfully report variant-associated phenotypes as compared to transgenic overexpression. Data for nine knock-in mutants across five genes are presented that demonstrate high editing efficiencies (60%), a reduced screening workload (24 F1 progeny), and a rapid timescale (4-5 d). This optimized method simplifies genome engineering and is readily adaptable to other model systems.