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. 2012 Jan 19;7(1):e30429. doi: 10.1371/journal.pone.0030429

Figure 5. PCR and WB of KLF8 after shRNA knockdown in U87-MG.

Figure 5

(A) U87-MG cells transfected with either scrambled (scr) or KLF8-shRNA (kd) were cultivated for up to 3 days (day 0 = day of seeding). Cells were harvested every 24 hrs and RNA was isolated, transcribed into cDNA and amplified by qPCR; data were normalized relative to levels of the house keeping gene TBP. Semi-quantitative qPCR displayed a clear knock-down in KLF8 expression already 48 hrs after transfection (day 0). Expression levels decreased to about 10% in KLF8-shRNA treated cells compared to cells treated with scrambled shRNA on day 2 after seeding. (B) Subsequent Western Blot analysis of the nuclear fraction of KLF8-kd U87-MG cells on day 4 after seeding revealed that KLF8 protein was still detectable in all transfected U87-MG cells but only to a small extent in the KLF8-knockdown cells indicating that shRNA-knockdown was successful in these transfected cells in concordance with the qPCR results (Figure 5A).