Construction and Evaluation of MKN45 DEC1 Knockout Gene Stable Strain by Using CRISPR/Cas9 Gene Editing System

Xiao-Li MA, Zhi-Tao CHEN, Shan-Hui SUN, Shu-Yi HAN, Yun-Shan WANG, Xiao-Hong SHI, Yan-Fei JIA

Abstract


Human differentiated embryonic chondrocyte-expressed gene 1 (DEC1) (also known as Stra13/Bhlhb2/Sharp2) is a basic helix-loop-helix (bHLH) transcription factor. Our previously studies have shown that the upregulation of DEC1 play an important role in cell proliferation in gastric cancer (GC). The latest gene editing system, CRISPR/Cas9 technology uses sequence-specific RNA molecules guide endonuclease Cas9 to the target nucleic acid and active the endonuclease to cut the double-stranded DNA, thereby complete genome knockout. Therefore, we use the CRISPR/ Cas9 system to completely knockout DEC1 genes in GC. Two gRNAs against DEC1 were designed. 293T cells were transfected using Lipofectamine 2000 and packaged for the recombinant lentivirus particles. Subsequently, we collected the lentivirus venom to infect the MKN45 cells and establish a stable, knockout cell line named MKN45-DEC1-KO. This study will provide a new cell model for further study of the role of DEC1 in the pathogenesis of gastric cancer.

Keywords


DEC1, CRISPR/Cas9, MKN45


DOI
10.12783/dtbh/mshh2017/13424

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