Systematic genome editing of the genes on zebrafish Chromosome 1 by CRISPR/Cas9
Systematic genome editing of the genes on zebrafish Chromosome 1 by CRISPR/Cas9
复制标题
通过 CRISPR/Cas9 对斑马鱼 1 号染色体上的基因进行系统基因组编辑
DOI：
10.1101/gr.248559.119
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发表时间：
2020-01-01
期刊：
影响因子：
7
通讯作者：
Sun, Jiawei
中科院分区：
文献类型：
--
作者：
Sun, Yonghua;Zhang, Bo;Sun, Jiawei
通过完善的CRISPR/Cas9技术进行基因组编辑，极大地促进了我们对许多生物过程的理解。然而，对任何物种或模式生物的完整全基因组敲除很少实现。在这里，我们对斑马鱼1号染色体上的所有基因（1333个）进行了系统敲除，成功地突变了1029个基因，并产生了1039个种系可传播的等位基因，对应于636个基因。同时，通过高通量生物信息学分析，我们发现序列特征在gRNA有效靶向感兴趣的特定基因中起着关键作用，而基因靶向的成功率与靶点GC含量呈正相关。此外，我们发现近四分之一的突变体与人类疾病有关，这里描述了几个具有代表性的CRISPR/ cas9产生的突变体。此外，我们试图确定导致基因突变体和反义morpholino介导的敲低胚胎之间不同表型的潜在机制。总之，这项工作通过CRISPR/Cas9技术生成了第一个全染色体范围的斑马鱼基因突变体集合，这将为社区提供宝贵的资源，我们的生物信息学分析也为设计基因特异性grna以成功进行基因编辑提供了一些有用的指导。
Genome editing by the well-established CRISPR/Cas9 technology has greatly facilitated our understanding of many biological processes. However, a complete whole-genome knockout for any species or model organism has rarely been achieved. Here, we performed a systematic knockout of all the genes (1333) on Chromosome 1 in zebrafish, successfully mutated 1029 genes, and generated 1039 germline-transmissible alleles corresponding to 636 genes. Meanwhile, by high-throughput bioinformatics analysis, we found that sequence features play pivotal roles in effective gRNA targeting at specific genes of interest, while the success rate of gene targeting positively correlates with GC content of the target sites. Moreover, we found that nearly one-fourth of all mutants are related to human diseases, and several representative CRISPR/Cas9-generated mutants are described here. Furthermore, we tried to identify the underlying mechanisms leading to distinct phenotypes between genetic mutants and antisense morpholino-mediated knockdown embryos. Altogether, this work has generated the first chromosome-wide collection of zebrafish genetic mutants by the CRISPR/Cas9 technology, which will serve as a valuable resource for the community, and our bioinformatics analysis also provides some useful guidance to design gene-specific gRNAs for successful gene editing.