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Construction of green fluorescent protein (GFP)‐marked strains of Bradyrhizobium for ecological studies
Author(s) -
Bhatia R.,
Dogra R.C.,
Sharma P.K.
Publication year - 2002
Publication title -
journal of applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.889
H-Index - 156
eISSN - 1365-2672
pISSN - 1364-5072
DOI - 10.1046/j.1365-2672.2002.01768.x
Subject(s) - green fluorescent protein , bradyrhizobium , vigna , biology , plasmid , microbial inoculant , bradyrhizobium japonicum , gene , microbiology and biotechnology , rhizobiaceae , botany , bacteria , genetics , symbiosis , rhizobium
Aim: To introduce the gfp gene encoding green fluorescent protein (GFP) into bradyrhizobia for their identification in nodules, soil and carrier‐based inoculants. Methods and Results:Bradyrhizobium sp. strains M29 and GN7, which nodulate mungbean ( Vigna radiata ), were conjugated with Escherichia coli S17‐1 carrying plasmid EDS 15 (a suicide plasmid carrying a promoterless gfp gene fused with Tn 5). The GFP‐marked strain expressed the gfp gene from a Bradyrhizobium promoter and gave green fluorescence when observed under an epifluorescent microscope or u.v. transilluminater. All the GFP‐marked strains were able to nodulate mungbean and fix nitrogen. The GFP‐marked bradyrhizobia were recovered at a frequency of 90–100% and 16–63% from nodules formed under sterilized and unsterilized conditions, respectively. The GFP‐marked bradyrhizobia were identified from soil and from charcoal‐based inoculants on the basis of green fluorescence. Conclusions: The GFP‐marked Bradyrhizobium was successfully identified on the basis of green fluorescence to study its competition and survival in the soil and in charcoal‐based inoculants. Significance and Impact of the Study: Introduction of the gfp gene into Bradyrhizobium provides a simple, specific and cost‐effective method of strain identification for ecological studies.