Prime editing in chicken fibroblasts and primordial germ cells
Prime editing in chicken fibroblasts and primordial germ cells
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DOI：
10.1111/dgd.12823
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发表时间：
2022-11-29
影响因子：
2.5
通讯作者：
Saito, Daisuke
中科院分区：
文献类型：
--
作者：
Atsuta, Yuji;Suzuki, Katsuya;Saito, Daisuke
基于CRISPR/Cas9的基因组编辑技术正在给发育生物学带来革命性的变化。基于CRISPR的先进技术之一是基序编辑(PE)，它能够在多个模式生物中进行精确的基因修改。然而，到目前为止，还没有关于利用PE系统在原始生殖细胞(PGCs)中进行基因编辑的报道。在目前的研究中，我们描述了一种将PE应用于鸡成纤维细胞和PGCs基因组的方法。通过将PE与转座子介导的基因组整合、药物选择和单细胞培养方法相结合，我们成功地产生了初始编辑的鸡成纤维细胞和PGCs。鸡PGC被广泛用于研究生殖细胞形成的实验模型和作为基因转移的载体来生产转基因鸡。这种实验模型在发育生物学领域很有用，也是生产药用和营养蛋白质的潜在生物反应器。因此，本文提出的方法不仅为研究生殖细胞发育中的基因功能提供了有力的工具，而且也为培育初级编辑的转基因禽类提供了基础。
CRISPR/Cas9-based genome editing technologies are revolutionizing developmental biology. One of the advanced CRISPR-based techniques is prime editing (PE), which enables precise gene modification in multiple model organisms. However, there has been no report of taking advantage of the PE system for gene editing in primordial germ cells (PGCs) thus far. In the current study, we describe a method to apply PE to the genome of chicken fibroblasts and PGCs. By combining PE with a transposon-mediated genomic integration, drug selection, and the single-cell culture method, we successfully generated prime-edited chicken fibroblasts and PGCs. The chicken PGC is widely used as an experimental model to study germ cell formation and as a vector for gene transfer to produce transgenic chickens. Such experimental models are useful in the developmental biology field and as potential bioreactors to produce pharmaceutical and nutritious proteins. Thus, the method presented here will provide not only a powerful tool to investigate gene function in germ cell development but also a basis for generating prime-edited transgenic birds.