Fig. 2
From: High-performance gene expression and knockout tools using sleeping beauty transposon system
Overexpression of FBW7 using various vectors and identification of new associated protein of FBW7 though Tandem Affinity Purification. a Map of pSB-SFB-FBW7 vector. SFB-tagged FBW7 protein is composed of S-Peptide, Flag, and Streptavidin-binding Peptide. b The expression of FBW7 and its downstream target cyclin E in HeLa cells were analyzed using Western blot with indicated antibodies. c Four HeLa cell lines expressing either Flag-FBW7, Myc-FBW7, HA-FBW7 or V5-FBW7 are indicated. HeLa cells were co-transfected with pSB-GFP-Flag/Myc/HA/V5-FBW7 and SB100X plasmids for 24 h, and then selected with puromycin for 72 h. d HeLa cells stably overexpressing SFB-FBW7 fusion protein analyzed by Western blot (n = 3). e Silver staining of the SFB-FBW7 complex in SDS-PAGE gel. The whole-cell extracts were prepared from HeLa cells, and the purification steps were as indicated (n = 3). f Identification of FBW7 interacting protein by mass spectrometry. FBW7-interacting proteins, including FBW7 and NFATc1, are indicated (n = 3)