Establishment of PC12 cell line with stable inhibition of DAPK expression / 基础医学与临床

ABSTRACT

Objective To screen a cell line which stably suppresses DAPK expression and to observe the growth characters.Methods Four pairs of shRNA were designed,synthesized and inserted into the pDsRed1-N1-U6 vector.The recombinant plasmids were purified and transfected into PC12 cell.Meanwhile,a pDsRed1-N1-U6 vector was transfected as control.The cell clones were screened by G418,and the stable PC12 cell line was established.DAPK expression was detected by Western blot.MTT method and Flow Cytometry(FCM) assay were used to assess the growth characters of the cell line.Results The shRNAs were transfected into PC12 cell and the cell clones were successfully screened out.Of the four recombinant plasmids,the F2 was the best interfering shRNA.Beyond our expectation,the F1 recombinant plasmid had an enhanced effect on DAPK expression.Conclusion A stable PC12 cell line with stable inhibition of DAPK expression by was established using siRNA expression vectors.