Project description:A shotgun microarray-based approach was used to identify candidate genes encoding the FOS utilization pathway in L. paracasei 1195. Differential expression profiles between cells grown on FOS and glucose provided a basis for identifying genes specifically induced by FOS. In addition, transcriptional analysis of cells grown on FOS or FOS + glucose allowed us to investigate the effect of catabolite repression on the expression of the FOS-induced genes. Keywords: gene identification
Project description:A shotgun microarray-based approach was used to identify candidate genes encoding the FOS utilization pathway in L. paracasei 1195. Differential expression profiles between cells grown on FOS and glucose provided a basis for identifying genes specifically induced by FOS. In addition, transcriptional analysis of cells grown on FOS or FOS + glucose allowed us to investigate the effect of catabolite repression on the expression of the FOS-induced genes. For sugar induction experiment (FOS vs. glucose; FOS vs. fructose), L. paracasei were grown in modified MRS (mMRS) basal medium to a final OD625 nm of ca. 0.3, at which point all of the residual sugars in the medium were consumed (based on HPLC analysis of the supernatants). The culture was then divided into equal portions, where FOS or glucose was added to a 1% final concentration. Cells were collected for total RNA isolation after 30 min (OD625 nm ca. 0.3 â 0.4). For glucose repression experiment (FOS vs. FOS + glucose), cells were grown in mMRS supplemented with 2% FOS to an OD625 nm 0.6. The culture was split into equal portions, and glucose was added to one of the portions at a 2% final concentration. Both cultures were grown for another 60 min (OD625 nm ca. 1.0) before harvested for total RNA isolation. Both experiments were performed in independent replicates (two biological replicates per experiment) with incubation at 37 degree celsius in ambient atmosphere.
