Fig. 1 Supplement

Fig. 1 Supplement Optimization of petroleum ether extraction and recoating. (A) Mean OD₆₀₀ values of control or delipidated M. marinum recovered in complete growth medium. (B) Mean mg of lipid extracted per gram of dry bacteria from initial bacteria and from delipidated bacteria following designated recovery time in culture. (C) Mean mg of lipid extracted per gram of dry bacteria following recoating with designated lipid densities. Two-tailed, unpaired t test. (D) Thin-layer chromatography (TLC) showing no difference between extracts from untreated or recoated bacteria. PDIM standard, petroleum ether extracts from untreated bacteria, and petroleum ether extracts from bacteria that were recoated with lipids from petroleum ether extracts (recoated). TLC was ran once in 98:2 petroleum ether:ethyl acetate. (E) 1 mg of extract from three separate petroleum ether extracts was dissolved in DMSO and separated by SDS-PAGE and stained with Colloidal Coomassie Brilliant Blue for 24 hr. Ladder is precision plus protein kaleidoscope. (F) TLC showing PDIM’s complete removal following petroleum ether extraction. PDIM standard, petroleum ether extract (PE lipids), chloroform:methanol extract of the bacterial pellet previously extracted with petroleum ether (PE-CHCL₃:CH₃OH lipids), and chloroform:methanol extract of a fresh bacterial pellet (CHCL₃:CH₃OH lipids). TLC was ran twice in 98:2 petroleum ether:ethyl acetate. (G) TLC of PDIM standard, PNDIM standard, and petroleum ether extracts of wildtype (WT) and ∆mmpL7 M. marinum. TLC was ran twice in 98:2 petroleum ether:ethyl acetate. (A)-(D) and (F)-(G) representative of three separate experiments.