小柯机器人

新型基因组编辑方法问世
2019-10-22 14:56

美国哈佛大学David R. Liu小组在最新研究中报道了一种不需要产生DNA双链断裂以及不需要供体DNA的基因组编辑方法。2019年10月21日,《自然》在线发表了相关论文。

研究人员描述了一个名为prime编辑(prime editing)的方法,这是一种通用且精确的基因组编辑方法,它使用融合了工程逆转录酶的催化受损的Cas9将新的遗传信息直接写入指定的DNA位点,并使用主要编辑向导RNA(pegRNA)进行编程,两者均指定了目标位点并编码所需的编辑。研究人员在人类细胞中进行了175次以上的编辑,包括靶向插入、缺失和所有12种类型的点突变,而无需双链断裂或供体DNA模板。

研究人员在人类细胞中应用了主要编辑功能,以有效地纠正镰状细胞疾病(需要HBB基因发生转化)和Tay-Sachs病(需要HEXA基因发生缺失)的主要遗传原因,并利用少量副产物在PRNP基因中加入保护性转化,并将各种标签和表位精确插入目标基因座。四个人类细胞系和原代有丝分裂后的小鼠皮层神经元以不同的效率支持prime编辑。

与碱基编辑相比,prime编辑在同源臂介导的修复、互补的优劣势等方面的效率和产物纯度方面提供了优势,并且在已知Cas9脱靶位点处的脱靶编辑比Cas9核酸酶低得多。prime编辑大大扩展了基因组编辑的范围和能力,并且原则上可以纠正约89%的已知致病性人类遗传变异。

据介绍,大多数导致疾病的遗传变异很难有效纠正,同时产生过多的副作用。

附:英文原文
 
Title:Search-and-replace genome editing without double-strand breaks or donor DNA
 
Author:Andrew V. Anzalone, Peyton B. Randolph, Jessie R. Davis, Alexander A. Sousa, Luke W. Koblan, Jonathan M. Levy, Peter J. Chen, Christopher Wilson, Gregory A. Newby, Aditya Raguram & David R. Liu
 
Issue&Volume:21 October 2019
 
Abstract: 
 
Most genetic variants that contribute to disease1 are challenging to correct efficiently and without excess byproducts2–5. Here we describe prime editing, a versatile and precise genome editing method that directly writes new genetic information into a specified DNA site using a catalytically impaired Cas9 fused to an engineered reverse transcriptase, programmed with a prime editing guide RNA (pegRNA) that both specifies the target site and encodes the desired edit. We performed more than 175 edits in human cells including targeted insertions, deletions, and all 12 types of point mutation without requiring double-strand breaks or donor DNA templates. We applied prime editing in human cells to correct efficiently and with few byproducts the primary genetic causes of sickle cell disease (requiring a transversion in HBB) and Tay-Sachs disease (requiring a deletion in HEXA), to install a protective transversion in PRNP, and to insert various tags and epitopes precisely into target loci. Four human cell lines and primary post-mitotic mouse cortical neurons support prime editing with varying efficiencies. Prime editing offers efficiency and product purity advantages over homology-directed repair, complementary strengths and weaknesses compared to base editing, and much lower off-target editing than Cas9 nuclease at known Cas9 off-target sites. Prime editing substantially expands the scope and capabilities of genome editing, and in principle could correct about 89% of known pathogenic human genetic variants.
 
DOI:10.1038/s41586-019-1711-4
 
Source: https://www.nature.com/articles/s41586-019-1711-4

Nature:《自然》,创刊于1869年。隶属于施普林格·自然出版集团,最新IF:69.504
官方网址:http://www.nature.com/
投稿链接:http://www.nature.com/authors/submit_manuscript.html


本期文章:《自然》:Online/在线发表

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