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Fig. 2 | BMC Veterinary Research

Fig. 2

From: Characterization of WWOX expression and function in canine mast cell tumors and malignant mast cell lines

Fig. 2

Generation of canine mast cell lines expressing WWOX lentiviral constructs or shRNAs targeting WWOX. a Full-length canine WWOX cDNA was PCR amplified from canine testes and cloned into the pCDH-copGFP lentiviral expression plasmid (Systems Biosciences). Canine BR cells transduced with empty control vector (EV) or WWOX lentivirus were sorted 72 h post-transduction to greater than 95% purity based on GFP expression. WWOX expression was assessed by qRT-PCR and Western blotting in empty vector or WWOX-expressing BR cells. b Canine C2 cells transduced with either scramble (Scr) control vector or pGreenPuro-shWWOX-copGFP lentiviral constructs (shWWOX-554 or shWWOX-1304) underwent FACs-mediated cell sorting based on GFP expression 72 h post-transduction. RNA and protein lysates were generated and qRT-PCR and Western blotting for WWOX and β-actin was performed to confirm efficiency of WWOX knockdown. Full-length blots are presented in Supplementary Fig. 1. Gene expression was calculated using the ΔCt method and all samples were normalized to 18S. Data represent mean +/− SD of three independent experiments and representative blots from three experimental replicates are shown. (Bars: SD. Statistical analysis: one-way ANOVA, *p ≤ 0.05, ***p ≤ 0.001)

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