Fig. 10

Characterization of C. reinhardtii DGAT3 expressed in E. coli. The coding sequence of C. reinhardtii dgat3 was cloned into pet19b vector (pet19b-DG) and transformed into E coli cells. The empty vector (pet19b) was transformed as a control. Protein induction was done on both DGAT3-expressing and control cells. Samples were taken before (t₀) and 4 h after induction (4 h). a Soluble proteins from both samples were analyzed by 12% SDS-PAGE and either stained with Coomassie brilliant blue (upper panel) or transferred to nitrocellulose (bottom panel). The membranes were incubated with rabbit anti-C. reinhardtii DGAT3 polyclonal antisera. The protein standards are shown on the right lanes, the dashes indicate the extra band in the gel upon induction and the protein recognized by the antisera, of approximately 40 kDa. b Lipids from E. coli cells were extracted with methanol: chloroform (1:1) and analyzed by thin layer chromatography (TLC) using a mix of hexane-diethyl ether-formic acid as the solvent system. Standards are: CE, chloesterol ester; W, wax ester; TAG, triglyceride; FFA, free fatty acids; Chol, cholesterol; PL, phospholipids