|Lorio, Julio - UNIV OF MISSOURI-COLUMBIA|
Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: March 6, 2000
Publication Date: N/A
Technical Abstract: The symbiosis between Sinorhizobium fredii USDA257 and soybean [Glycine max (L.) Merr.] exhibits a high degree of cultivar specificity. Sinorhizobium fredii USDA257 nodulates 'primitive' soybean but fails to nodulate the improved cultivar McCall. The present work provides evidence for the involvement of a novel genetic locus in controlling cultivar specificity. The new locus was identified in S. fredii USDA257 by Tn5 transposon mutagenesis followed by nodulation screening on different hosts. Sinorhizobium fredii USDA257GCV is a Tn5 mutant that nodulates the soybean cultivar McCall. Southern hybridization analysis demonstrated that the new genetic locus was not located in the region corresponding to the previously identified plasmid-borne cultivar specificity, no1XWBTUV operon. The DNA sequence of the cloned region tagged by the Tn5 insertion displayed three open-reading frames (ORFs) with high degrees of identity to the gcv operon in Escherichia coli and other organisms. In E. coli the gcv operon encodes three proteins with apparent molecular masses of 39 kD (T-protein), 17.3 kD (H-protein), and 90 kD (P-protein). The ORF disrupted by the Tn5 transposon in S. fredii USDA257GCV codes for the aminomethyltransferase precursor (T-protein). Use of SDS/PAGE with in vivo **35S-methionine labeled proteins showed that the mutant lacks a polypeptide corresponding to the molecular mass of the P-protein (90 kDa). A cosmid containing a wild-type gcv region was mobilized into the mutant background causing restoration of P-protein synthesis. Currently, we are determining the role of the gcv operon in cultivar-specific nodulation of S. fredii USDA257.