%0 Journal Article
%T Construction of EZH2 Knockout Animal Model by CRISPR/Cas9 Technology
%A Fanrong MENG
%A Dan ZHAO
%A Qinghua ZHOU
%A Zhe LIU
%J Öйú·Î°©ÔÓÖ¾
%D 2018
%R : 10.3779/j.issn.1009-3419.2018.05.02
%X Background and objective It has been proven that CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats/CRISPR-associated 9) system was the modern gene-editing technology through the constitutive expression of nucleases Cas9 in the mammalian, which binds to the specific site in the genome mediated by single-guide RNA (sgRNA) at desired genomic loci. The aim of this study is that the animal model of EZH2 gene knockout was constructed using CRISPR/Cas9 technology. Methods In this study, we designed two single-guide RNAs targeting the Exon3 and Exon4 of EZH2 gene. Then, their gene-targeting efficiency were detected by SURVEYOR assay. The lentivirus was perfused into the lungs of mice by using a bronchial tube and detected by immunohistochemistry and qRT-PCR. Results The experimental results of NIH-3T3 cells verify that the designed sgEZH2 can efficiently effect the cleavage of target DNA by Cas9 in vitro. The immunohistochemistry and qRT-PCR results showed that the EZH2 expression in experimental group was significantly decreased in the mouse lung tissue. Conclusion The study successfully designed two sgRNA which can play a knock-out EZH2 function. An EZH2 knockout animal model was successfully constructed by CRISPR/Cas9 system, and it will be an effective animal model for studying the functions and mechanisms of EZH2.£¿
%K CRISPR-Cas9
%K EZH2
%K sgRNA
%K Gene knockout
%U http://www.lungca.org/index.php?journal=01&page=article&op=view&path%5B%5D=10.3779%2Fj.issn.1009-3419.2018.05.02