Effect Of Interference With E2F1 Gene Expression On Proliferation And Migration Of EBV-Positive Lymphoma Raji Cells

[Objective] E2 F transcription factor 1(E2F1)is overexpressed in a variety of human tumors.Our previous experiments also found that E2F1 gene is highly expressed in EBV-transformed lymphoblasts(LCLs).In this experiment,Raji cells stably expressing E2F1 gene were constructed,and the expression of E2F1 gene was detected by q RT-PCR and Western blot.The proliferation and migration of Raji cells after interference with E2F1 gene expression were observed by CCK-8 assay and migration invasion assay.To observe the effects of E2F1 gene expression on the proliferation and migration of EBV-positive lymphoma Raji cells.[Method] q RT-PCR and Western Blot were used to detect the expression of E2F1 gene in EBV-transformed lymphoblasts(LCL7)and EBV-positive Raji cells,and the lentiviral vector E2F1-sh RNA-GFP-PURO against E2F1 gene was constructed to establish E2F1 stability.The low-expression Raji cell line(Raji-E2F1-sh RNA1,Raji-E2F1-sh RNA2)was observed by inverted fluorescence microscopy,and the interference effect of E2F1 gene was verified by q RT-PCR and Western Blot.The effects of interference E2F1 gene expression on proliferation and migration of EBV-positive lymphoma Raji cell line were observed by CCK-8 assay and migration invasion assay.[Results] 1.The expression level of E2F1 in EBV-positive lymphoma Raji cells is higher than that of EBV-transformed lymphoblasts(LCL7).In this study,the expression of E2F1 gene in Raji cells and LCL7 cells was detected by q RT-PCR and Western blot.The expression of E2F1 gene in Raji was higher than that in LCL7(P<0.01).Tips Raji cells can be used for subsequent experiments.2.Successful establishment of a stable Raji cell line with low expression of E2F1 geneThe results of plasmid sequencing showed that the interference plasmid E2F1-sh RNA1-GFP-PURO of the target gene was completely consistent with the designed interference sequence by Genomic sequencing,indicating that the vector was successfully constructed.The green fluorescence of the transfected group and the group was observed by inverted fluorescence microscope.Carrying rate,up to 80%;compare the E2F1 interference efficiency of Raji/E2F1 sh RNA1,Raji/E2F1 sh RNA2 transfected with E2F1-sh RNA-GFP-PURO plasmid by q RT-PCR and Western Blot,and select Raji/E2F1 with higher interference efficiency.sh RNA1 was inserted into the subsequent experiments;after comparison,q RT-PCR and Western Blot results showed that the expression level of E2F1 in cells transfected with E2F1-sh RNA1-GFP-PURO plasmid(interference group,Raji/E2F1 sh RNA1)was higher than that of transfected Raji-SH-GFP-The cells of the PURO plasmid(empty vector group,Raji/NC)and common Raji cells(blank group)were significantly reduced,suggesting that the interference effect was good,and the subsequent experiments could be continued.3.Interfering with E2F1 gene expression can inhibit the proliferation and invasion of EBV-positive lymphoma cellsThe results of CCK-8 showed that the proliferation of Raji/NC sh RNA1 in the interference group was significantly slower than that in the empty vector group(P<0.05),indicating that the interference of E2F1 gene expression could inhibit the proliferation of Raji cells.The results of soft agar colony formation showed that the interference ability of Raji/E2F1 sh RNA1 in the interference group was lower than that in the empty vector group(P<0.05).The cell cycle results of flow cytometry showed that the proportion of Raji/E2F1 sh RNA1 in the interference group was significantly higher than that in the empty vector group Raji/NC,and the percentage of G1 and G2 cells decreased,the difference was statistically significant(P<0.05).Interference with E2F1 gene expression can lead to S phase arrest in lymphoma Raji cells.The results of migration experiments showed that the interference group Raji/E2F1 sh RNA1 was significantly smaller than the empty vector group Raji/NC,and the difference was statistically significant(P<0.05),indicating that the interference of E2F1 gene expression can significantly reduce the migration ability of lymphoma Raji cells..The results of invasion assay showed that the number of cells in the interference group Raji/E2F1 sh RNA1 crossing Matrigel Matrigel was significantly lower than that in the empty vector Raji/NC,the difference was statistically significant(P<0.05),indicating that interference with E2F1 gene expression can inhibit lymphoma Raji cells.Invasive ability.Conclusion: Interfering with E2F1 gene expression can inhibit the proliferation and migration of EBV-positive lymphoma Raji cells.