siRNA Transfection Assay for Centrosomal Proteins

Transfections of siRNA were performed using Lipofectamine RNAiMAX transfection reagent (Thermo Fisher Scientific) according to the manufacturer’s instructions. 30,000–40,000 cells were seeded per well in a 24-well plate and analyzed 72 h after the initial transfection. The following siRNAs (50 nM final concentration) were used: siLuciferase (siControl), 5′-UCG​AAG​UAU​UCC​GCG​UAC​G-3′ (Dharmacon ON-TARGET plus/Ambion Silencer Select; Knodler et al., 2010 (link)); siNek2, 5′-GAU​GCA​AUU​UGG​UCA​UUA​AUU-3′ and 5′-GAA​AGG​CAA​UAC​UUA​GAU​GUU-3′ (Dharmacon ON-TARGET plus; Kim et al., 2015 (link)); siCep164, 5′-GGU​GAC​AUU​UAC​UAU​UUC​A-3′ (Ambion Silencer Select; Graser et al., 2007 (link)); siHsODF2-1, 5′-AGA​CUA​AUG​GAG​CAA​CAA​G-3′ (Ambion Silencer Select; Soung et al., 2009 (link)); siODF2-2, 5′-GGA​UCU​UUA​UGU​CGC​UGA​ATT-3′ (Ambion Silencer Select); siKif24, 5′-GGA​ACA​CCC​TGG​AGA​ATA​GTT-3′ and 5′-GAG​TTG​AGC​TCT​CCT​TTG​GTT (Dharmacon ON-TARGET plus; Kobayashi et al., 2011 (link)); and siC-Nap1 (human CEP250), 5′-GAG​CAG​AGC​UAC​AGC​GAA​U-3′ and 5′-AAG​CUG​ACG​UGG​UGA​AUA​A-3′ (Dharmacon ON-TARGET plus; Panic et al., 2015 (link)).