Generating a CRISPR knockout mouse through a strong premature termination codon: a cautionary tale
Qing Rex Lyu1
Peng Yao2
Joseph M.Miano3
1.Aab Cardiovascular Research Institute, University of Rochester School of Medicine &Dentistry,Rochester, NY 14642,USA;Vascular Biology Center,Medical College of Georgia at Augusta University,Augusta, GA 30912,USA;Institute of Biomedicine and Health,Chongqing Institute of Green and Intelligent Technology,Chinese Academy of Sciences,Chongqing 400714,China2.Aab Cardiovascular Research Institute, University of Rochester School of Medicine &Dentistry,Rochester, NY 14642,USA3.Aab Cardiovascular Research Institute, University of Rochester School of Medicine &Dentistry,Rochester, NY 14642,USA;Vascular Biology Center,Medical College of Georgia at Augusta University,Augusta, GA 30912,USA
摘要：Dear Editor,
Genetic inactivation or gene knockout (KO) has been a powerful method of elucidating gene function in mice.The traditional method of generating KO mice via targeting mouse embryonic stem cells through homologous recombination is labor-intensive and of low efficiency.The emergence of clustered regularly interspaced short palindromic repeat (CRISPR) technology greatly simplifies the method of gene targeting at a reduced cost and with high efficiency.In its simplest form, CRISPR editing comprises two components, a single guide RNA (sgRNA) and Cas9 protein.Upon finding the complementary sequence preceding a protospacer adjacent motif (PAM), this ribonucleoprotein (RNP) complex generates a double-strand break (DSB) three nucleotides upstream of the PAM.The DSB is then repaired via error-prone non-homologous end-joining (NHEJ).Alternatively, a third component may be used involving a single-strand or double-strand repair template engineered with nucleotide substitutions, deletions or insertions of new genetic material[1].The presence of such repair templates biases the reparation of the DSB via homology directed repair (HDR) processes.
机标关键词：
分类号：Q789(基因工程（遗传工程）)
论文发表日期：2021-03-30
在线出版日期：2025-08-15（本平台首次上网日期，不代表文献的发表时间）
页数：5( 174-178 )