Abstract Drought stress is the main limiting factor of soybean yield. Currently, genetic engineering has been one important tool in the development of drought-tolerant cultivars. A widely used strategy is the fusion of genes that confer tolerance under the control of the CaMV35S constitutive promoter; however, stress-responsive promoters would constitute the best alternative to the generation of drought-tolerant crops. We characterized the promoter of α-galactosidase soybean (GlymaGAL) gene that was previously identified as highly up-regulated by drought stress. The β-glucuronidase (GUS) activity of Arabidopsis transgenic plants bearing 1000- and 2000-bp fragments of the GlymaGAL promoter fused to the uidA gene was evaluated under air-dried, polyethylene glycol (PEG) and salt stress treatments. After 24 h of air-dried and PEG treatments, the pGAL-2kb led to an increase in GUS expression in leaf and root samples when compared to the control samples. These results were corroborated by qPCR expression analysis of the uidA gene. The pGAL-1kb showed no difference in GUS activity between control and treated samples. The pGAL-2kb promoter was evaluated in transgenic soybean roots, leading to an increase in EGFP expression under air-dried treatment. Our data indicates that pGAL-2kb could be a useful tool in developing drought-tolerant cultivars by driving gene expression.

Metabolomics analysis of wild type Arabidopsis thaliana plants, under control and drought stress conditions revealed several metabolic pathways that are induced under water deficit. The metabolic response to drought stress is also associated with ABA dependent and independent pathways, allowing a better understanding of the molecular mechanisms in this model plant. Through combining an in silico approach and gene expression analysis by quantitative real-time PCR, the present work aims at identifying genes of soybean metabolic pathways potentially associated with water deficit. Digital expression patterns of Arabidopsis genes, which were selected based on the basis of literature reports, were evaluated under drought stress condition by Genevestigator. Genes that showed strong induction under drought stress were selected and used as bait to identify orthologs in the soybean genome. This allowed us to select 354 genes of putative soybean orthologs of 79 Arabidopsis genes belonging to 38 distinct metabolic pathways. The expression pattern of the selected genes was verified in the subtractive libraries available in the GENOSOJA project. Subsequently, 13 genes from different metabolic pathways were selected for validation by qPCR experiments. The expression of six genes was validated in plants undergoing drought stress in both pot-based and hydroponic cultivation systems. The results suggest that the metabolic response to drought stress is conserved in Arabidopsis and soybean plants.

The study of tolerance mechanisms for drought stress in soybean is fundamental to the understanding and development of tolerant varieties. Using in silico analysis, four marker genes involved in the classical ABA-dependent and ABA-independent pathways of drought response were identified in the Glycine max genome in the present work. The expression profiles of the marker genes ERD1-like, GmaxRD20A-like, GmaxRD22-like and GmaxRD29B-like were investigated by qPCR in root samples of drought sensitive and tolerant soybean cultivars (BR 16 and Embrapa 48, respectively), submitted to water deficit conditions in hydroponic and pot-based systems. Among the four putative soybean homologs to Arabidopsis genes investigated herein, only GmaxRD29B-like was not regulated by water deficit stress. Distinct expression profiles and different induction levels were observed among the genes, as well as between the two drought-inducing systems. Our results showed contrasting gene expression responses for the GmaxRD20A-like and GmaxRD22-like genes. GmaxRD20A-like was highly induced by continuous drought acclimating conditions, whereas GmaxRD22-like responses decreased after abrupt water deprivation. GmaxERD1-like showed a different expression profile for the cultivars in each system. Conversely, GmaxRD20A-like and GmaxRD22-like genes exhibited similar expression levels in tolerant plants in both systems.

The neuraminidase gene, nanH, is present in the O1, non-toxigenic Vibrio cholerae Amazonia strain. Its location has been assigned to a 150 kb NotI DNA fragment, with the use of pulsed-field gel electrophoresis and DNA hybridization. This NotI fragment is positioned inside 630 kb SfiI and 1900 kb I-CeuI fragments of chromosome 1. Association of the pathogenicity island VPI-2, carrying nanH and other genes, with toxigenic strains has been described by other authors. The presence of nanH in a non-toxigenic strain is an exception to this rule. The Amazonia strain nanH was sequenced (Genbank accession No. AY825932) and compared to available V. cholerae sequences. The sequence is different from those of pandemic strains, with 72 nucleotide substitutions. This is the first description of an O1 strain with a different nanH allele. The most variable domain of the Amazonia NanH is the second lectin wing, comprising 13 out of 17 amino acid substitutions. Based on the presence of nanH in the same region of the genome, and similarity of the adjacent sequences to VPI-2 sequences, it is proposed that the pathogenicity island VPI-2 is present in this strain.