CRISPR-Cas9 Expressed in Stably Transduced Cell Lines Promotes Recombination and Selects for Herpes Simplex Virus Recombinants.
CRISPR-Cas9 Expressed in Stably Transduced Cell Lines Promotes Recombination and Selects for Herpes Simplex Virus Recombinants.
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DOI：
10.1016/j.crviro.2022.100023
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发表时间：
2022-06
期刊：
影响因子：
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通讯作者：
H. Oh;F. Diaz;Changhong Zhou;Nicholas Carpenter;D. Knipe
中科院分区：
文献类型：
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作者：
H. Oh;F. Diaz;Changhong Zhou;Nicholas Carpenter;D. Knipe
重组单纯疱疹病毒的构建有多种方法，包括同源重组、细菌人工染色体操作和酵母遗传方法。同源重组可能具有在病毒基因组中引入较少基因改变的优势，但如果没有筛选或选择，重组体的低水平可能会使这种方法更加耗时。在这项研究中，我们使用了互补的表达Cas9的细胞系，并引导针对亲本病毒的RNA快速产生重组病毒。对子代病毒的分析表明，CRISPR-Cas9在促进重组以增加重组病毒的同时，对转染子病毒中的亲本病毒具有选择性。该方法也可用于丰富由任何当前方法制成的重组体。
Recombinant herpes simplex virus strains can be constructed by several methods, including homologous recombination, bacterial artificial chromosome manipulation, and yeast genetic methods. Homologous recombination may have the advantage of introducing fewer genetic alterations in the viral genome, but the low level of recombinants can make this method more time consuming if there is no screen or selection. In this study we used complementing cell lines that express Cas9 and guide RNAs targeting the parental virus to rapidly generate recombinant viruses. Analysis of the progeny viruses indicated that CRISPR-Cas9 both promoted recombination to increase recombinant viruses and selected against parental viruses in the transfection progeny viruses. This approach can also be used to enrich for recombinants made by any of the current methods.