Fig. 4
Validation of shRNA-GFP construct transfection into CAL27 cells and MK2 knockdown. a For the validation of MK2KD the expression levels of MK2 were confirmed by Western blot analysis against non-transfected controls using MK2-specific antibody. Negligible expression in shRNA 2 and shRNA Mix transfected cells as compared to normal and scrambled controls confirmed that the protein expression of MK2 has been down-regulated in these. C=Non-transfected control; 1, 2, 3, 4, SC = shRNA-GFP construct 1, 2, 3, 4, Scrambled Control transfected, respectively; Mix = Co-transfected with shRNA-GFP constructs 1, 2, 3, 4. GFP served as an input control while β tubulin was used as a loading control in this case. b qRT-PCR analysis of the CAL27-MK2KD cells established that the shRNA transfection lead to ~ 80% knockdown of MK2 compared to non-transfected control as shown in the histograms. All the qRT-PCR assays were performed atleast thrice. The results are expressed as means±standard errors of the mean