title: Isolation of a gramicidin S hyperproducing strain of Bacillus brevis by use of a fluorescence activated cell sorting system.
abstract: A gramicidin S (GS) hyperproducing mutant of Bacillus brevis was isolated by using a protein-staining fluorescence dye (fluorescein isothiocyanate, FITC), and a fluorescence-activated cell sorting system (FACS). By flow cytometry (FCM) analysis after staining with FITC, higher producing cells of the wild-type had higher fluorescence signals than cells with low productivity or cells from a GS non-producing mutant. Staining with FITC did not affect the viability of cells under the conditions chosen for FCM analysis. This enabled us to recover viable cells after sorting. After wild-type cells were mutagenized with N-methyl-N'-nitro-N-nitrosoguanidine, mutants with higher fluorescence than the parental strain were obtained by cell sorting. Among them, strain 18 was chosen as a GS hyperproducer; it produced 590 micrograms GS/ml compared to 350 micrograms/ml by the wild-type strain. This method has the advantage of being able to screen large numbers of cells in a short time. Furthermore, use of the fluorescence dye technique will expand the use of FACS to the improvement of other cultures that produce metabolites that do not have a specific fluorescence or strong enough fluorescence for normal cell sorting.
authors.affiliation: Department of Biology, Massachusetts Institute of Technology, Cambridge 02139.
authors.forename: T
authors.initials: T
authors.lastname: Azuma
authors.forename: G I
authors.initials: GI
authors.lastname: Harrison
authors.forename: A L
authors.initials: AL
authors.lastname: Demain
mesh_terms: D001407:Bacillus / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D005434:Flow Cytometry; D016650:Fluorescein-5-isothiocyanate; D006096:Gramicidin / Q000096:biosynthesis*; D009154:Mutation; D013194:Staining and Labeling
chemical_list: D006096:Gramicidin; D016650:Fluorescein-5-isothiocyanate