Anti goat or anti rabbit alkaline phosphatase conjugated antibody

Western blot was performed as previously described [22] using the following primary antibodies: Caspase-3 and UVRAG (Cell Signaling Technology, Danvers, MA, USA) as well as KRIT1 and β-actin (Santa Cruz Biotechnology, Dallas, TX, USA). Anti-goat or anti-rabbit alkaline phosphatase-conjugated antibody (Promega, Madison, WI, USA) was used as a secondary antibody.
Target prediction and Luciferase activity assay miR-1185 target genes were predicted with TargetScan 6.2 (http://www.targetscan.org/). The pmiR-RB-REPORT TM dual luciferase reporter vectors carrying the wild-type (WT) or mutated (MUT) 3'-UTR of miR-1185 target genes (UVRAG and KRIT1) were constructed (Ribio Co., Guangzhou, China). Human embryonic kidney 293 (HEK-293) cells (ATCC, Manassas, VA, USA) were co-transfected with 200 ng of recombinant plasmid, 50 nM of miR-1185 mimic and 100 nM of miR-1185 inhibitor. After 24 h of transfection, the luciferase activities were measured with a dual luciferase reporter assay kit (Promega, Madison, WI, USA) on a luminometer (GloMaxTM 20/20, Promega).