In soybean, the W4 gene encoding dihydroflavonol-4-reductase controls anthocyanin pigment biosynthesis in flowers. The mutant allele, w4-m, is characterized by variegated flowers and was evolved from the insertion of an endogenous transposable element, Tgm9, in intron II of the W4 gene. In the w4-m mutant line, reversion of the unstable allele from variegated to normal purple flower in revertants would indicate Tgm9's excision accompanied by its insertion into a second locus. We identified a male-sterile, female-sterile mutant from such germinal revertant bearing purple flowers. The objectives of our investigation were to map the sterility locus, identify candidate genes for the male-fertile, female-fertile phenotype, and then determine if sterility is associated with the insertion of Tgm9 in the sterility locus. We used bulked segregant analysis to map the locus to molecular linkage group J (chromosome 16). Fine mapping enabled us to flank the locus to a 62-kb region that contains only five predicted genes. One of the genes in that region, Glyma16g07850.1, codes for a helicase. A rice homolog of this gene has been shown to control crossing over and fertility phenotype. Thus, Glyma16g07850.1 is most likely the gene regulating the male and female fertility phenotype in soybean. DNA blot analysis of the segregating individuals for Tgm9 showed perfect association between sterility and the presence of the transposon. Most likely, the sterility mutation was caused by the insertion of Tgm9. The transposable element should facilitate identification of the male- and female-fertility gene. Characterization of the fertility gene will provide vital molecular insight on the reproductive biology of soybean and other plants.

The W4 locus in soybean encodes a dihydroflavonol-4-reductase (DFR2) that regulates pigmentation patterns in flowers and hypocotyl. The mutable w4-m allele that governs variegated flowers has arisen through insertion of a CACTA-type transposable element, Tgm9, in DFR2. In the w4-m line, reversion from variegated to purple flower indicates excision of Tgm9, and its insertion at a new locus. Previously, we have identified a male-sterile, female-sterile mutant among the selfed progenies of a revertant plant carrying only purple flowers. Co-segregation between Tgm9 and the sterility phenotype suggested that the mutant was generated by insertion of Tgm9 at the St8 locus. The transposon was localized to exon 10 of Glyma.16G072300 that shows high identity to the MER3 DNA helicase involved in crossing over. Molecular analysis of fertile branches from two independent revertant plants confirmed precise excision of Tgm9 from the st8 allele, which restored fertility. In soybean, the gene is expressed in flower-buds, trifoliate leaves and stem. Phylogenetic analysis placed St8 in a clade with the Arabidopsis and rice MER3 suggesting that St8 is most likely the orthologous MER3 soybean gene. This study established the utility of Tgm9 in gene identification as well as in forward and reverse genetics studies.

Sudden death syndrome (SDS) is caused by the fungal pathogen, Fusarium virguliforme, and is a major threat to soybean production in North America. There are two major components of this disease: (i) root necrosis and (ii) foliar SDS. Root symptoms consist of root necrosis with vascular discoloration. Foliar SDS is characterized by interveinal chlorosis and leaf necrosis, and in severe cases by flower and pod abscission. A major toxin involved in initiating foliar SDS has been identified. Nothing is known about how root necrosis develops. In order to unravel the mechanisms used by the pathogen to cause root necrosis, the transcriptome of the pathogen in infected soybean root tissues of a susceptible cultivar, ‘Essex’, was investigated. The transcriptomes of the germinating conidia and mycelia were also examined. Of the 14,845 predicted F. virguliforme genes, we observed that 12,017 (81%) were expressed in germinating conidia and 12,208 (82%) in mycelia and 10,626 (72%) in infected soybean roots. Of the 10,626 genes induced in infected roots, 224 were transcribed only following infection. Expression of several infection-induced genes encoding enzymes with oxidation-reduction properties suggests that degradation of antimicrobial compounds such as the phytoalexin, glyceollin, could be important in early stages of the root tissue infection. Enzymes with hydrolytic and catalytic activities could play an important role in establishing the necrotrophic phase. The expression of a large number of genes encoding enzymes with catalytic and hydrolytic activities during the late infection stages suggests that cell wall degradation could be involved in root necrosis and the establishment of the necrotrophic phase in this pathogen.

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