BackgroundThe entire world is facing a deteriorating environment. Understanding the mechanisms underlying plant responses to external abiotic stresses is important for breeding stress-tolerant crops and herbages. Phytohormones play critical regulatory roles in plants in the response to external and internal cues to regulate growth and development. Medicago falcata is one of the stress-tolerant candidate leguminous species and is able to fix atmospheric nitrogen. This ability allows leguminous plants to grow in nitrogen deficient soils.MethodsWe performed Illumina sequencing of cDNA prepared from abiotic stress treated M. falcata. Sequencedreads were assembled to provide a transcriptome resource. Transcripts were annotated using BLASTsearches against the NCBI non-redundant database and gene ontology definitions were assigned. Acomparison among the three abiotic stress treated samples was carried out. The expression of transcriptswas confirmed with qRT-PCR.ResultsWe present an abiotic stress-responsive M. falcata transcriptome using next-generation sequencing data from samples grown under standard, dehydration, high salinity, and cold conditions. We combined reads from all samples and de novo assembled 98,515 transcripts to build the M. falcata gene index. A comprehensive analysis of the transcriptome revealed abiotic stress-responsive mechanisms underlying the metabolism and core signalling components of major phytohormones. We identified nod factor signalling pathways during early symbiotic nodulation that are modified by abiotic stresses. Additionally, a global comparison of homology between the M. falcata and M. truncatula transcriptomes, along with five other leguminous species, revealed a high level of global sequence conservation within the family.ConclusionsM. falcata is shown to be a model candidate for studying abiotic stress-responsive mechanisms in legumes. This global gene expression analysis provides new insights into the biochemical and molecular mechanisms involved in the acclimation to abiotic stresses. Our data provides many gene candidates that might be used for herbage and crop breeding. Additionally, FalcataBase (http://bioinformatics.cau.edu.cn/falcata/) was built for storing these data.Electronic supplementary materialThe online version of this article (doi:10.1186/s12864-015-2019-x) contains supplementary material, which is available to authorized users.

Background: Alfalfa is a high-quality forage cultivated widely in northern China. Recently, the failure of alfalfa plants to survive the winter has caused substantial economic losses. Water management has attracted considerable attention as a method for the potential improvement of winter survival. The aim of this study was to determine whether and how changes in the water regime affect the freezing tolerance of alfalfa. Results: The alfalfa variety WL353LH was cultivated under water regimes of 80 and 25% of water-holding capacity, and all the plants were subjected to low temperatures at 4/0°C (light/dark) and then − 2/− 6°C (light/dark). The semi-lethal temperatures were lower for water-stressed than well-watered alfalfa. The pool sizes of total soluble sugars, total amino acids, and proline changed substantially under water-deficit and low-temperature conditions. Metabolomics analyses revealed 72 subclasses of differential metabolites, among which lipid and lipid-like molecules (e.g., fatty acids, unsaturated fatty acids, and glycerophospholipids) and amino acids, peptides, and analogues (e.g., proline betaine) were upregulated under water-deficit conditions. Some carbohydrates (e.g., Dmaltose and raffinose) and flavonoids were also upregulated at low temperatures. Finally, Kyoto Encyclopedia of Genes and Genomes analyses revealed 18 significantly enriched pathways involved in the biosynthesis and metabolism of carbohydrates, unsaturated fatty acids, amino acids, and glycerophospholipids. Conclusions: Water deficit significantly enhanced the alfalfa' freezing tolerance, and this was correlated with increased soluble sugar, amino acid, and lipid and lipid-like molecule contents. These substances are involved in osmotic regulation, cryoprotection, and the synthesis, fluidity, and stability of the cellular membrane. Our study provides a reference for improving alfalfa' winter survival through water management.

Alfalfa, an important forage legume, is an ideal crop for sustainable agriculture and a potential crop for bioenergy resources. Drought, one of the most common environmental stresses, substantially affects plant growth, development, and productivity. MicroRNAs (miRNAs) are newly discovered gene expression regulators that have been linked to several plant stress responses. To elucidate the role of miRNAs in drought stress regulation of alfalfa, a high-throughput sequencing approach was used to analyze 12 small RNA libraries comprising of four samples, each with three biological replicates. From the 12 libraries, we identified 348 known miRNAs belonging to 80 miRNA families, and 281 novel miRNAs, using Mireap software. Eighteen known miRNAs in roots and 12 known miRNAs in leaves were screened as drought-responsive miRNAs. With the exception of miR319d and miR157a which were upregulated under drought stress, the expression pattern of drought-responsive miRNAs was different between roots and leaves in alfalfa. This is the first study that has identified miR3512, miR3630, miR5213, miR5294, miR5368 and miR6173 as drought-responsive miRNAs. Target transcripts of drought-responsive miRNAs were computationally predicted. All 447 target genes for the known miRNAs were predicted using an online tool. This study provides a significant insight on understanding drought-responsive mechanisms of alfalfa.

The induction of miR399 and miR398 and the inhibition of miR156, miR159, miR160, miR171, miR2111, and miR2643 were observed under Pi deficiency in alfalfa. The miRNA-mediated genes involved in basic metabolic process, root and shoot development, stress response and Pi uptake. Inorganic phosphate (Pi) deficiency is known to be a limiting factor in plant development and growth. However, the underlying miRNAs associated with the Pi deficiency-responsive mechanism in alfalfa are unclear. To elucidate the molecular mechanism at the miRNA level, we constructed four small RNA (sRNA) libraries from the roots and shoots of alfalfa grown under normal or Pi-deficient conditions. In the present study, alfalfa plants showed reductions in biomass, photosynthesis, and Pi content and increases in their root-to-shoot ratio and citric, malic, and succinic acid contents under Pi limitation. Sequencing results identified 47 and 44 differentially expressed miRNAs in the roots and shoots, respectively. Furthermore, 909 potential target genes were predicted, and some targets were validated by RLM-RACE assays. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses showed prominent enrichment in signal transducer activity, binding and basic metabolic pathways for carbohydrates, fatty acids and amino acids; cellular response to hormone stimulus and response to auxin pathways were also enriched. qPCR results verified that the differentially expressed miRNA profile was consistent with sequencing results, and putative target genes exhibited opposite expression patterns. In this study, the miRNAs associated with the response to Pi limitation in alfalfa were identified. In addition, there was an enrichment of miRNA-targeted genes involved in biological regulatory processes such as basic metabolic pathways, root and shoot development, stress response, Pi transportation and citric acid secretion.