Construction of eukaryotic vector BMI-1 shRNA and its inhibition in Hela cells

Objective:To construct eukaryotic vector expressing short hairpin RNA(shRNA)of BMI-1,and to inhibit BMI-1 mRNA expression in human cervix cancer cell line Hela.Method:Designing four different shRNA targeting the coding sequence of BMI-1,the BMI-1 shRNA1,BMI-1 shRNA2,BMI-1 shRNA3 and BMI-1 shRNA4 were constructed by inserting the designed shRNA to the eukaryotic expression vector psiRNA-hH1neo,and transfected into Hela cells with Lipofectamine 2000.Result:It was verified by partial nucleotide sequencing that the constructed eukaryotic vectors expressing shRNA of BMI-1 were correct.The BMI-1 expression of mRNA was significantly suppressed by 59.9% and 67.4% in BMI-1 shRNA 2 and 4 transfected Hela cells as compared with untransfected Hela cells,respectively.Conclusion:The vector of BMI-1 shRNA 1,BMI-1 shRNA 2,BMI-1 shRNA3 and BMI-1 shRNA4 has been successfully constructed.BMI-1 shRNA 2 and BMI-1 shRNA 4 could efficiently suppress BMI-1 expression in Hela cells.This work has laid foundations for further study on cancer gene therapy and potential application of BMI-1.