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Figure 4 | Breast Cancer Research

Figure 4

From: Enhanced RAD21 cohesin expression confers poor prognosis and resistance to chemotherapy in high grade luminal, basal and HER2 breast cancers

Figure 4

Validation of RAD21 expression in MDA-MB-231 cells carrying shRAD21 knockdown constructs. A. Quantitative real time PCR analysis of RAD21 expression in clones stably expressing two different shRAD21 constructs and shRNAmir vector, relative to the parental cell line. B. Western blot analysis of RAD21 protein level. RAD21 expression in five independently derived clonal cell lines with two different stable RAD21 knockdown constructs was compared to the parental cell line, cells transfected with shRNAmir vector and an immortalized human breast epithelial cell line MCF10A. Pan-actin was used as loading control. A reduction in the levels of RAD21 protein in three stable clones (sh223_sc1, sh224_sc4 and sh224_sc5) was verified by semi-quantitative Western blot analysis (bottom panel). The levels of RAD21 protein were normalized to the pan-actin loading control and expressed as the percentage of the parental line where the RAD21 expression was given an arbitrary value of 100%. The values represent the mean of four independent experiments except for shRNAmir vector and MCF10A where three independent experiments were performed. Clones with a significant reduction in either RAD21 mRNA or protein levels were marked by asterisks (* P < 0.05; ** P < 0.005, Student t-test). Error bar = SEM.

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