A CRISPR-Cas9 Assisted Non-Homologous End-Joining Strategy for One-step Engineering of Bacterial Genome
A CRISPR-Cas9 Assisted Non-Homologous End-Joining Strategy for One-step Engineering of Bacterial Genome
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DOI：
10.1038/srep37895
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发表时间：
2016-11-24
影响因子：
4.6
通讯作者：
Qi, Qingsheng
中科院分区：
文献类型：
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作者：
Su, Tianyuan;Liu, Fapeng;Qi, Qingsheng
同源重组介导的基因组工程已在原核生物中得到广泛应用，具有高效、准确的特点。然而，由于DNA编辑模板构建过程相对复杂，该方法在实现大量基因或大DNA片段的更大规模基因组编辑方面受到限制。在这里，我们描述了一种 CRISPR-Cas9 辅助的非同源末端连接 (CA-NHEJ) 策略，以不依赖同源重组的方式快速有效地灭活细菌基因，且无需使用选择性标记。我们的研究表明，CA-NHEJ 可用于一步删除大的染色体 DNA 片段，不需要同源 DNA 模板。因此，它是减少细菌基因组的一种新颖而强大的工具，并具有加速基因组进化的潜力。
Homologous recombination-mediated genome engineering has been broadly applied in prokaryotes with high efficiency and accuracy. However, this method is limited in realizing larger-scale genome editing with numerous genes or large DNA fragments because of the relatively complicated procedure for DNA editing template construction. Here, we describe a CRISPR-Cas9 assisted non-homologous end-joining (CA-NHEJ) strategy for the rapid and efficient inactivation of bacterial gene (s) in a homologous recombination-independent manner and without the use of selective marker. Our study show that CA-NHEJ can be used to delete large chromosomal DNA fragments in a single step that does not require homologous DNA template. It is thus a novel and powerful tool for bacterial genomes reducing and possesses the potential for accelerating the genome evolution.