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. 2012 Apr 12;7(4):e34440. doi: 10.1371/journal.pone.0034440

Figure 2. Expression of NOX5 in RCSC determined by RT-PCR and western blot.

Figure 2

(A) NOX5 expression detected by RT-PCR using human NOX5-specific PCR primers. The amplified product was confirmed as a NOX5 sequence by nucleotide sequencing (n = 6). (B) Knockdown of the steady state levels of NOX5 mRNA. Values obtained by densitometry were analyzed and ratios were calculated. Amount of knockdown of NOX5 mRNA steady pools are depicted as vertical bars. (1) siNTC (40 nM), (2) siNOX5 (5 nM), (3) siNOX5 (10 nM), (4) siNOX5 (20 nM), (5) siNOX5 (40 nM). (C) Western blot analysis detected by NOX5 specific antibody. RCS cells were grown, harvested, lysed, and fractionated by centrifugation at 29,000× g. The pellets were suspended in lysis buffer and subjected to IP for enriching the NOX5 followed by Western blot analysis. Controls included NOX5 encoding HUVEC and anti NOX5 blocking peptide (NOX5 BP). (D) Knockdown of NOX5 determined by Western blotting. RCSC were transfected with 40 nM non target control siRNA (siNTC) or siRNA against NOX5 (siNOX5). After 72 h, the cells were harvested and total protein lysates were used for Western blot analysis by NOX5 specific antibody. β-Actin was used as loading control.