RISPR/Cas9方法构建神经丛素B1敲除的Caco-2结直肠癌细胞株

he RNA-guided Cas9 nuclease from the microbial clustered regularly interspaced short palindromic repeat sequences（CRISPR）has been used to facilitate efficient genome engineering. The CRISPR/Cas9 technology also has been adapted into working of gene knockout in eukaryotic cells. Plexin-B1, as Sema4D receptor, plays an important role in tumor angiogenesis. To further investigate the function of plexin-B1, Caco-2 colorectal cell line were selected as research model for Plexin-B1 gene knockout by CRISPR/Cas9 system. CRISPR/Cas9-Plexin-B1 plasmids were constructed, then were transfected into Caco-2 cells. Plexin-B1-/- Caco-2 cell strain were selected through cell screening. The results showed that Plexin-B1 gene could be successfully knockout in Caco-2 cells by CRISPR/Cas9 technology. Plexin-B1-/- Caco-2 cell strain were then screened out. Phosphorylation of PI3K(a important signaling molecule in tumor angiogenesis) were reduced in pelxin-B1 knockout caco-2 cells.