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The ability to respond quickly to salt stress can determine the tolerance level of a species. Here, we test how rapidly the roots of Calotropis procera react to high salinity conditions. In the first 24 h after saline exposure, the plants reduced stomatal conductance, increased CO(2) assimilation, and water use efficiency. Thus, the root tissue showed an immediate increase in soluble sugars, free amino acid, and soluble protein contents. Twelve aquaporins showed differential gene expression in the roots of C. procera under salinity. Transcriptional upregulation was observed only after 2 h, with greater induction of CpTIP1.4 (fourfold). Transcriptional downregulation, in turn, occurred mainly after 8 h, with the largest associated with CpPIP1.2 (fourfold). C. procera plants responded quickly to high saline levels. Our results showed a strong stomatal control associated with high free amino acid and soluble sugar contents, regulated aquaporin expression in roots, and supported the high performance of the root system of C. procera under salinity. Moreover, this species was able to maintain a lower Na(+)/K(+) ratio in the leaves compared to that of the roots of stressed plants. The first response of the root system, after immediate contact with saline solution, present an interesting scenario to discuss. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12298-021-00957-9.

Cenostigma pyramidale is a native legume of the Brazilian semiarid region which performs symbiotic association with arbuscular mycorrhizal fungi (AMF), being an excellent model for studying genes associated with tolerance against abiotic and biotic stresses. In RT-qPCR approach, the use of reference genes is mandatory to avoid incorrect interpretation of the relative expression. This study evaluated the stability of ten candidate reference genes (CRGs) from C. pyramidale root tissues under salt stress (three collection times) and associated with AMF (three different times of salinity). The de novo transcriptome was obtained via RNA-Seq sequencing. Three algorithms were used to calculate the stability of CRGs under different conditions: (i) global (Salt, Salt+AMF, AMF and Control, and collection times), (ii) only non-inoculated plants, and (iii) AMF (only inoculated plants). HAG2, SAC1, aRP3 were the most stable CRGs for global and AMF assays, whereas HAG2, SAC1, RHS1 were the best for salt stress assay. This CRGs were used to validate the relative expression of two up-regulated transcripts in Salt2h (RAP2-3 and PIN8). Our study provides the first set of reference genes for C. pyramidale under salinity and AMF, supporting future researches on gene expression with this species.

Even before the perception or interaction with pathogens, plants rely on constitutively guardian molecules, often specific to tissue or stage, with further expression after contact with the pathogen. These guardians include small molecules as antimicrobial peptides (AMPs), generally cysteine-rich, functioning to prevent pathogen establishment. Some of these AMPs are shared among eukaryotes (eg, defensins and cyclotides), others are plant specific (eg, snakins), while some are specific to certain plant families (such as heveins). When compared with other organisms, plants tend to present a higher amount of AMP isoforms due to gene duplications or polyploidy, an occurrence possibly also associated with the sessile habit of plants, which prevents them from evading biotic and environmental stresses. Therefore, plants arise as a rich resource for new AMPs. As these molecules are difficult to retrieve from databases using simple sequence alignments, a description of their characteristics and in silico (bioinformatics) approaches used to retrieve them is provided, considering resources and databases available. The possibilities and applications based on tools versus database approaches are considerable and have been so far underestimated.

Calotropis procera is a perennial Asian shrub with significant adaptation to adverse climate conditions and poor soils. Given its increased salt and drought stress tolerance, C. procera stands out as a powerful candidate to provide alternative genetic resources for biotechnological approaches. The qPCR (real-time quantitative polymerase chain reaction), widely recognized among the most accurate methods for quantifying gene expression, demands suitable reference genes (RGs) to avoid over- or underestimations of the relative expression and incorrect interpretation. This study aimed at evaluating the stability of ten RGs for normalization of gene expression of root and leaf of C. procera under different salt stress conditions and different collection times. The selected RGs were used on expression analysis of three target genes. Three independent experiments were carried out in greenhouse with young plants: i) Leaf100 = leaf samples collected 30 min, 2 h, 8 h and 45 days after NaCl-stress (100 mM NaCl); ii) Root50 and iii) Root200 = root samples collected 30 min, 2 h, 8 h and 1day after NaCl-stress (50 and 200 mM NaCl, respectively). Stability rank among the three algorithms used showed high agreement for the four most stable RGs. The four most stable RGs showed high congruence among all combination of collection time, for each software studied, with minor disagreements. CYP23 was the best RG (rank of top four) for all experimental conditions (Leaf100, Root50, and Root200). Using appropriated RGs, we validated the relative expression level of three differentially expressed target genes (NAC78, CNBL4, and ND1) in Leaf100 and Root200 samples. This study provides the first selection of stable reference genes for C. procera under salinity. Our results emphasize the need for caution when evaluating the stability RGs under different amplitude of variable factors.

Phytopathogenic microorganisms have a significant influence on survival and productivity of several crop plants. Transcription factors (TFs) are important players in the response to biotic stresses, as insect attack and pathogen infection. In face of such adversities many TFs families have been previously reported as differentially expressed in plants as a reaction to bacterial, fungal and viral infection. This review highlights recent progresses in understanding the structure, function, signal regulation and interaction of transcription factors with other proteins in response to pathogens. Hence, we focus on three families of transcription factors: ERF, bZIP and WRKY, due to their abundance, importance and the availability of functionally well-characterized members in response to pathogen attack. Their roles and the possibilities related to the use of this knowledge for engineering pathogen resistance in crop plants are also discussed.

Plants exhibit sensitive mechanisms to respond to environmental stresses, presenting some specific and non-specific reactions when attacked by pathogens, including organisms from different classes and complexity, as viroids, viruses, bacteria, fungi and nematodes. A crucial step to define the fate of the plant facing an invading pathogen is the activation of a compatible Resistance (R) gene, the focus of the present review. Different aspects regarding R-genes and their products are discussed, including pathogen recognition mechanisms, signaling and effects on induced and constitutive defense processes, splicing and post transcriptional mechanisms involved. There are still countless challenges to the complete understanding of the mechanisms involving R-genes in plants, in particular those related to the interactions with other genes of the pathogen and of the host itself, their regulation, acting mechanisms at transcriptional and post-transcriptional levels, as well as the influence of other types of stress over their regulation. A magnification of knowledge is expected when considering the novel information from the omics and systems biology.

Plants have the ability to recognize and respond to a multitude of pathogens, resulting in a massive reprogramming of the plant to activate defense responses including Resistance (R) and Pathogenesis-Related (PR) genes. Abiotic stresses can also activate PR genes and enhance pathogen resistance, representing valuable genes for breeding purposes. The present work offers an overview of soybean R and PR genes present in the GENOSOJA (Brazilian Soybean Genome Consortium) platform, regarding their structure, abundance, evolution and role in the plant-pathogen metabolic pathway, as compared with Medicago and Arabidopsis. Searches revealed 3,065 R candidates (756 in Soybean, 1,142 in Medicago and 1,167 in Arabidopsis), and PR candidates matching to 1,261 sequences (310, 585 and 366 for the three species, respectively). The identified transcripts were also evaluated regarding their expression pattern in 65 libraries, showing prevalence in seeds and developing tissues. Upon consulting the SuperSAGE libraries, 1,072 R and 481 PR tags were identified in association with the different libraries. Multiple alignments were generated for Xa21 and PR-2 genes, allowing inferences about their evolution. The results revealed interesting insights regarding the variability and complexity of defense genes in soybean, as compared with Medicago and Arabidopsis.

Physical maps are important tools to uncover general chromosome structure as well as to compare different plant lineages and species, helping to elucidate genome structure, evolution and possibilities regarding synteny and colinearity. The increasing production of sequence data has opened an opportunity to link information from mapping studies to the underlying sequences. Genome browsers are invaluable platforms that provide access to these sequences, including tools for genome analysis, allowing the integration of multivariate information, and thus aiding to explain the emergence of complex genomes. The present work presents a tutorial regarding the use of genome browsers to develop targeted physical mapping, providing also a general overview and examples about the possibilities regarding the use of Fluorescent In Situ Hybridization (FISH) using bacterial artificial chromosomes (BAC), simple sequence repeats (SSR) and rDNA probes, highlighting the potential of such studies for map integration and comparative genetics. As a case study, the available genome of soybean was accessed to show how the physical and in silico distribution of such sequences may be compared at different levels. Such evaluations may also be complemented by the identification of sequences beyond the detection level of cytological methods, here using members of the aquaporin gene family as an example. The proposed approach highlights the complementation power of the combination of molecular cytogenetics and computational approaches for the anchoring of coding or repetitive sequences in plant genomes using available genome browsers, helping in the determination of sequence location, arrangement and number of repeats, and also filling gaps found in computational pseudochromosome assemblies.

Physical maps are important tools to uncover general chromosome structure as well as to compare different plant lineages and species, helping to elucidate genome structure, evolution and possibilities regarding synteny and colinearity. The increasing production of sequence data has opened an opportunity to link information from mapping studies to the underlying sequences. Genome browsers are invaluable platforms that provide access to these sequences, including tools for genome analysis, allowing the integration of multivariate information, and thus aiding to explain the emergence of complex genomes. The present work presents a tutorial regarding the use of genome browsers to develop targeted physical mapping, providing also a general overview and examples about the possibilities regarding the use of Fluorescent In Situ Hybridization (FISH) using bacterial artificial chromosomes (BAC), simple sequence repeats (SSR) and rDNA probes, highlighting the potential of such studies for map integration and comparative genetics. As a case study, the available genome of soybean was accessed to show how the physical and in silico distribution of such sequences may be compared at different levels. Such evaluations may also be complemented by the identification of sequences beyond the detection level of cytological methods, here using members of the aquaporin gene family as an example. The proposed approach highlights the complementation power of the combination of molecular cytogenetics and computational approaches for the anchoring of coding or repetitive sequences in plant genomes using available genome browsers, helping in the determination of sequence location, arrangement and number of repeats, and also filling gaps found in computational pseudochromosome assemblies.