利用CRISPR/Cas9基因编辑技术构建Nup155基因敲除的PK-15细胞系OA北大核心

To study the effect of Nup155 function on viral replication,CRISPR/Cas9 gene editing technology was used to construct Nup155 knockout PK-15 cell lines.Four gRNAs were designed to precisely target the Nup155 gene.The sgRNAs were synthesized and then fused with the LentiCRISPR v2 vector.The sgRNA vectors were packaged as a lentivirus in 293 cells.Cell lines were screened with 2 μg/mL purinomycin.RT-qPCR and Western-blot were used to confirm whether Nup155 gene was knocked out in cells.Finally,CCK8 assay was used to detect the effect of Nup155 gene knockout on the activity of PK-15 cells.Both RT-qPCR and Western-blot results showed that the mRNA and protein expression levels of Nup155 in single cell clones were significantly reduced.CCK8 assay results showed that PK-15 cell viability decreased in Nup155 knocked out cell lines.In conclusion,this study successfully constructed a PK-15 cell line with Nup155 partial knocked out by CRISPR/Cas9 technology,and partial knocking out Nup155 significantly affected the viability of PK-15 cells.This study provides a useful cell model for further in-depth study of Nup155 function.