Chromosome-level genome assembly provides insights into the genome evolution and functional importance of the phenylpropanoid-flavonoid pathway in Thymus mongolicus.

BACKGROUND: Thymus mongolicus (family Lamiaceae) is a Thyme subshrub with strong aroma and remarkable environmental adaptability. Limited genomic information limits the use of this plant. RESULTS: Chromosome-level 605.2 Mb genome of T. mongolicus was generated, with 96.28% anchored to 12 pseudochromosomes. The repetitive sequences were dominant, accounting for 70.98%, and 32,593 protein-coding genes were predicted. Synteny analysis revealed that Lamiaceae species generally underwent two rounds of whole genome duplication; moreover, species-specific genome duplication was identified. A recent LTR retrotransposon burst and tandem duplication might play important roles in the formation of the Thymus genome. Using comparative genomic analysis, phylogenetic tree of seven Lamiaceae species was constructed, which revealed that Thyme plants evolved recently in the family. Under the phylogenetic framework, we performed functional enrichment analysis of the genes on nodes that contained the most gene duplication events (> 50% support) and of relevant significant expanded gene families. These genes were highly associated with environmental adaptation and biosynthesis of secondary metabolites. Combined transcriptome and metabolome analyses revealed that Peroxidases, Hydroxycinnamoyl-CoA shikimate/quinate hydroxycinnamoyl transferases, and 4-coumarate-CoA ligases genes were the essential regulators of the phenylpropanoid-flavonoid pathway. Their catalytic products (e.g., apigenin, naringenin chalcone, and several apigenin-related compounds) might be responsible for the environmental tolerance and aromatic properties of T. mongolicus. CONCLUSION: This study enhanced the understanding of the genomic evolution of T. mongolicus, enabling further exploration of its unique traits and applications, and contributed to the understanding of Lamiaceae genomics and evolutionary biology.

The Potential Role of Genic-SSRs in Driving Ecological Adaptation Diversity in Caragana Plants.

Caragana, a xerophytic shrub genus widely distributed in northern China, exhibits distinctive geographical substitution patterns and ecological adaptation diversity. This study employed transcriptome sequencing technology to investigate 12 Caragana species, aiming to explore genic-SSR variations in the Caragana transcriptome and identify their role as a driving force for environmental adaptation within the genus. A total of 3666 polymorphic genic-SSRs were identified across different species. The impact of these variations on the expression of related genes was analyzed, revealing a significant linear correlation (p < 0.05) between the length variation of 264 polymorphic genic-SSRs and the expression of associated genes. Additionally, 2424 polymorphic genic-SSRs were located in differentially expressed genes among Caragana species. Through weighted gene co-expression network analysis, the expressions of these genes were correlated with 19 climatic factors and 16 plant functional traits in various habitats. This approach facilitated the identification of biological processes associated with habitat adaptations in the studied Caragana species. Fifty-five core genes related to functional traits and climatic factors were identified, including various transcription factors such as MYB, TCP, ARF, and structural proteins like HSP90, elongation factor TS, and HECT. The roles of these genes in the ecological adaptation diversity of Caragana were discussed. Our study identified specific genomic components and genes in Caragana plants responsive to heterogeneous habitats. The results contribute to advancements in the molecular understanding of their ecological adaptation, lay a foundation for the conservation and development of Caragana germplasm resources, and provide a scientific basis for plant adaptation to global climate change.

One-step synthesis of phospho-rich, silica-enhanced chitosan aerogel for the efficient adsorption of uranium(VI).

In this study, an amorphous silica reinforced, phosphoric-crosslinked chitosan foam (P-CTS@SixOy) was prepared. The introduction of amorphous silica not only increased the affinity of the adsorbent for uranium, but also improved the stability of the material. The number of active sites of P-CTS@SixOy was increased by the introduction of phosphate groups. The material exhibited excellent uranium adsorption performance with the removal capacity and efficiency of 850.5 mg g-1 and 98.1 %, respectively. After regenerations, the morphology of P-CTS@SixOy still maintained, and the uranium adsorption efficiency remained above 90 %, manifesting the excellent cycle performance of P-CTS@SixOy. In the dynamic adsorption experiment, P-CTS@SixOy successfully concentrated the volume of uranium-containing solution, and exhibited excellent uranium adsorption performance. The analysis of kinetics, isotherms, and thermodynamics manifested that the uranium adsorption behavior of P-CTS@SixOy was a spontaneous, endothermic, monolayer chemical adsorption process. X-ray photoelectron spectroscopy, Scanning Electron Microscope, and Fourier Transform Infrared Spectrometer were used to characterized the P-CTS@SixOy before and after adsorption, which demonstrated that the main interaction mechanism between uranium and P-CTS@SixOy was the complexation. These studies indicated the huge application prospect of P-CTS@SixOy in the treatment of large-scale uranium-containing wastewater.

RADseq-based population genomic analysis and environmental adaptation of rare and endangered recretohalophyte Reaumuria trigyna.

Genetic diversity reflects the survival potential, history, and population dynamics of an organism. It underlies the adaptive potential of populations and their response to environmental change. Reaumuria trigyna is an endemic species in the Eastern Alxa and West Ordos desert regions in China. The species has been considered a good candidate to explore the unique survival strategies of plants that inhabit this area. In this study, we performed population genomic analyses based on restriction-site associated DNA sequencing to understand the genetic diversity, population genetic structure, and differentiation of the species. Analyses of 92,719 high-quality single-nucleotide polymorphisms (SNPs) indicated that overall genetic diversity of R. trigyna was low (HO = 0.249 and HE = 0.208). No significant genetic differentiation was observed among the investigated populations. However, a subtle population genetic structure was detected. We suggest that this might be explained by adaptive diversification reinforced by the geographical isolation of populations. Overall, 3513 outlier SNPs were located in 243 gene-coding sequences in the R. trigyna transcriptome. Potential sites under diversifying selection occurred in genes (e.g., AP2/EREBP, E3 ubiquitin-protein ligase, FLS, and 4CL) related to phytohormone regulation and synthesis of secondary metabolites which have roles in adaptation of species. Our genetic analyses provide scientific criteria for evaluating the evolutionary capacity of R. trigyna and the discovery of unique adaptions. Our findings extend knowledge of refugia, environmental adaption, and evolution of germplasm resources that survive in the Ordos area.

Elucidation of the evolutionary history of Stipa in China using comparative transcriptomic analysis.

Phylogenetic analysis provides crucial insights into the evolutionary relationships and diversification patterns within specific taxonomic groups. In this study, we aimed to identify the phylogenetic relationships and explore the evolutionary history of Stipa using transcriptomic data. Samples of 12 Stipa species were collected from the Qinghai-Tibet Plateau and Mongolian Plateau, where they are widely distributed, and transcriptome sequencing was performed using their fresh spikelet tissues. Using bidirectional best BLAST analysis, we identified two sets of one-to-one orthologous genes shared between Brachypodium distachyon and the 12 Stipa species (9397 and 2300 sequences, respectively), as well as 62 single-copy orthologous genes. Concatenation methods were used to construct a robust phylogenetic tree for Stipa, and molecular dating was used to estimate divergence times. Our results indicated that Stipa originated during the Pliocene. In approximately 0.8 million years, it diverged into two major clades each consisting of native species from the Mongolian Plateau and the Qinghai-Tibet Plateau, respectively. The evolution of Stipa was closely associated with the development of northern grassland landscapes. Important external factors such as global cooling during the Pleistocene, changes in monsoonal circulation, and tectonic movements contributed to the diversification of Stipa. This study provided a highly supported phylogenetic framework for understanding the evolution of the Stipa genus in China and insights into its diversification patterns.

Plant and Soil Microbial Diversity Co-Regulate Ecosystem Multifunctionality during Desertification in a Temperate Grassland.

Biodiversity plays a crucial role in driving multiple ecosystem functions in temperate grasslands. However, our understanding of how biodiversity regulates the impacts of desertification processes on ecosystem multifunctionality (EMF) remains limited. In this study, we investigate plant diversity, soil microbial diversity (fungal, bacterial, archaeal, and arbuscular mycorrhizal fungal (AMF) diversity), soil properties (soil water content, pH, and soil clay content), and multiple ecosystem functions (soil N mineralization, soil phosphatase activity, AMF infection rate, microbial biomass, plant biomass, and soil C and nutrients (N, P, K, Ca, Fe, Na, Cu, Mg, and Mn)) at six different grassland desertification intensities. The random forest model was conducted to assess the importance of soil properties, plant diversity, and soil microbial diversity in driving EMF. Furthermore, a structural equation model (SEM) was employed to analyze the indirect and direct impacts of these predictors on EMF. Our study showed that plant, soil bacterial, fungal, and archaeal diversity gradually decreased with increasing desertification intensity. However, only AMF diversity was found to be less sensitive to desertification. Similarly, EMF also showed a significant decline with increasing desertification. Importantly, both plant and soil microbial diversity were positively associated with EMF during desertification processes. The random forest model and SEM revealed that both plant and soil microbial diversity were identified as important and direct predictors of EMF during desertification processes. This highlights the primary influence of above- and below-ground biodiversity in co-regulating the response of EMF to grassland desertification. These findings have important implications for planned ecosystem restoration and sustainable grassland management.

Genomic insights into genetic diversity and local adaptation of a dominant desert steppe feather grass, Stipa breviflora Griseb.

Investigating the genetic mechanisms of local adaptation is critical to understanding how species adapt to heterogeneous environments. In the present study, we analyzed restriction site-associated DNA sequencing (RADseq) data in order to explore genetic diversity, genetic structure, genetic differentiation, and local adaptation of Stipa breviflora. In total, 135 individual plants were sequenced and 25,786 polymorphic loci were obtained. We found low genetic diversity (He = 0.1284) within populations of S. breviflora. Four genetic clusters were identified along its distribution range. The Mantel test, partial Mantel test, and multiple matrix regression with randomization (MMRR) indicate that population differentiation was caused by both geographic distance and environmental factors. Through the FST outlier test and environmental association analysis (EAA), 113 candidate loci were identified as putatively adaptive loci. RPK2 and CPRF1, which are associated with meristem maintenance and light responsiveness, respectively, were annotated. To explore the effects of climatic factors on genetic differentiation and local adaptation of S. breviflora, gradient forest (GF) analysis was applied to 25,786 single nucleotide polymorphisms (SNPs) and 113 candidate loci, respectively. The results showed that both temperature and precipitation affected the genetic differentiation of S. breviflora, and precipitation was strongly related to local adaptation. Our study provides a theoretical basis for understanding the local adaptation of S. breviflora.

Transcriptome-Wide Gene Expression Plasticity in Stipa grandis in Response to Grazing Intensity Differences.

Organisms have evolved effective and distinct adaptive strategies to survive. Stipa grandis is a representative species for studying the grazing effect on typical steppe plants in the Inner Mongolia Plateau. Although phenotypic (morphological and physiological) variations in S. grandis in response to long-term grazing have been identified, the molecular mechanisms underlying adaptations and plastic responses remain largely unknown. Here, we performed a transcriptomic analysis to investigate changes in gene expression of S. grandis under four different grazing intensities. As a result, a total of 2357 differentially expressed genes (DEGs) were identified among the tested grazing intensities, suggesting long-term grazing resulted in gene expression plasticity that affected diverse biological processes and metabolic pathways in S. grandis. DEGs were identified in RNA-Seq and qRT-PCR analyses that indicated the modulation of the Calvin-Benson cycle and photorespiration metabolic pathways. The key gene expression profiles encoding various proteins (e.g., ribulose-1,5-bisphosphate carboxylase/oxygenase, fructose-1,6-bisphosphate aldolase, glycolate oxidase, etc.) involved in these pathways suggest that they may synergistically respond to grazing to increase the resilience and stress tolerance of S. grandis. Our findings provide scientific clues for improving grassland use and protection and identifying important questions to address in future transcriptome studies.

Effect of arbuscular mycorrhizal symbiosis on ion homeostasis and salt tolerance-related gene expression in halophyte Suaeda salsa under salt treatments.

Halophytes can remove large quantities of salts from saline soils, so their importance in ecology has received increasing attention. Preliminary studies have shown that arbuscular mycorrhizal (AM) fungi can improve the salt tolerance of halophytes. However, few studies have focused on the molecular mechanisms and effects of AM fungi in halophytes under different salt conditions. A pot experiment was carried out to investigate the effects of Funneliformis mosseae inoculation on growth, nutrient uptake, ion homeostasis and the expression of salt tolerance-related genes in Suaeda salsa under 0, 100, 200 and 400 mM NaCl. The results showed that F. mosseae promoted the growth of S. salsa and increased the shoot Ca2+ and Mg2+ concentrations under no-salt condition and high-salt condition. In addition, AM fungi increased the K+ concentration and maintained a high K+/Na+ ratio at 400 mM NaCl, while AM fungi decreased the K+ concentration and reduced the K+/Na+ ratio at 0 mM NaCl. AM fungi downregulated the expression of SsNHX1 in shoots and the expression of SsSOS1 in roots at 400 mM NaCl. These effects may decrease the compartmentation of Na+ into leaf vacuoles and restrict Na+ transport from roots to shoots, leading to an increase in root Na+ concentration. AM symbiosis upregulated the expression of SsSOS1 in shoots and downregulated the expression of SsSOS1 and SsNHX1 in roots at 100 mM NaCl. However, regulation of the genes (SsNHX1, SsSOS, SsVHA-B and SsPIP) was not significantly different with AM symbiosis at 0 mM or 200 mM NaCl. The results revealed that AM symbiosis might induce diverse modulation strategies in S. salsa, depending on external Na+ concentrations. These findings suggest that AM fungi may play significant ecological roles in the phytoremediation of salinized ecosystems.

Climatic humidity mediates the strength of the species richness-biomass relationship on the Mongolian Plateau steppe.

The relationships between biodiversity and ecosystem functioning (BEF) have been extensively studied over past decades. However, the environmental factors affecting their relationships, and how their relationships vary under the influence of environmental factors, remain controversial. Studying the BEF relationships in natural/wild environments is of great significance for devising strategies in biodiversity conservation and ecosystem functioning. Using the data from 75 sites on the Mongolian Plateau steppe, we analyzed the relationship between species richness and biomass with general linear models (GLMs) and linear mixed models (LMMs), and analyzed the variation in the species richness-biomass relationships under environmental conditions by the partial least square path modeling (PLSPM). The results showed that de Martonne aridity index affected both species richness and community biomass in parallel, and that hydrothermal coupling conditions were more important direct impact factors for aboveground biomass. However, the significant species richness-biomass relationships became weaker when the effects of environmental factors (i.e. climate and soil properties) were present. Climate humidity was the most important factor in mediating the relationship between species richness and community biomass. Our research suggested that species richness-biomass relationships are weak in the natural grasslands of the Mongolian Plateau, and that this may be due to the differences in the regional-scale environment and changes in species interactions. We recommend that a more comprehensive understanding of the relationship between diversity and biomass requires further research within broader environmental gradients.

Identification of Genic SSRs Provide a Perspective for Studying Environmental Adaptation in the Endemic Shrub Tetraena mongolica.

Tetraena mongolica is a xerophytic shrub endemic to desert regions in Inner Mongolia. This species has evolved distinct survival strategies that allow it to adapt to hyper-drought and heterogeneous habitats. Simple sequence repeats (SSRs) may provide a molecular basis in plants for fast adaptation to environmental change. Thus, identifying SSRs and their possible effects on gene behavior has the potential to provide valuable information for studies of adaptation. In this study, we sequenced six individual transcriptomes of T. mongolica from heterogeneous habitats, focused on SSRs located in genes, and identified 811 polymorphic SSRs. Of the identified SSRs, 172, 470, and 76 were located in 5' UTRs, CDSs, and 3' UTRs in 591 transcripts; and AG/CT, AAC/GTT, and AT/AT were the most abundant repeats in each gene region. Functional annotation showed that many of the identified polymorphic SSRs were in genes that were enriched in several GO terms and KEGG pathways, suggesting the functional significance of these genes in the environmental adaptation process. The identification of polymorphic genic SSRs in our study lays a foundation for future studies investigating the contribution of SSRs to regulation of genes in natural populations of T. mongolica and their importance for adaptive evolution of this species.

Genome-wide identification and characterization of long non-coding RNAs involved in fruit ripening and the climacteric in Cucumis melo.

BACKGROUND: Cucumis melo is a suitable study material for investigation of fruit ripening owing to its climacteric nature. Long non-coding RNAs have been linked to many important biological processes, such as fruit ripening, flowering time regulation, and abiotic stress responses in plants. However, knowledge of the regulatory roles of lncRNAs underlying the ripening process in C. melo are largely unknown. In this study the complete transcriptome of Cucumis melo L. cv. Hetao fruit at four developmental stages was sequenced and analyzed. The potential role of lncRNAs was predicted based on the function of differentially expressed target genes and correlated genes. RESULTS: In total, 3857 lncRNAs were assembled and annotated, of which 1601 were differentially expressed between developmental stages. The target genes of these lncRNAs and the regulatory relationship (cis- or trans-acting) were predicted. The target genes were enriched with GO terms for biological process, such as response to auxin stimulus and hormone biosynthetic process. Enriched KEGG pathways included plant hormone signal transduction and carotenoid biosynthesis. Co-expression network construction showed that LNC_002345 and LNC_000154, which were highly expressed, might co-regulate with mutiple genes associated with auxin signal transduction and acted in the same pathways. We identified lncRNAs (LNC_000987, LNC_000693, LNC_001323, LNC_003610, LNC_001263 and LNC_003380) that were correlated with fruit ripening and the climacteric, and may participate in the regulation of ethylene biosynthesis and metabolism and the ABA signaling pathway. A number of crucial transcription factors, such as ERFs, WRKY70, NAC56, and NAC72, may also play important roles in the regulation of fruit ripening in C. melo. CONCLUSIONS: Our results predict the regulatory functions of the lncRNAs during melon fruit development and ripening, and 142 highly expressed lncRNAs (average FPKM > 100) were identified. These lncRNAs participate in the regulation of auxin signal transduction, ethylene, sucrose biosynthesis and metabolism, the ABA signaling pathway, and transcription factors, thus regulating fruit development and ripening.

Metagenome Profiling Identifies Potential Biocontrol Agents for Selaginella kraussiana in New Zealand.

Metagenomics can be used to identify potential biocontrol agents for invasive species and was used here to identify candidate species for biocontrol of an invasive club moss in New Zealand. Profiles were obtained for Selaginella kraussiana collected from nine geographically disjunct locations in Northern New Zealand. These profiles were distinct from those obtained for the exotic club moss Selaginella moellendorffii and the native club mosses Lycopodium deuterodensum and Lycopodium volubile also collected in Northern New Zealand. Fungi and bacteria implicated elsewhere in causing plant disease were identified on plants of Selaginella that exhibited signs of necrosis. Most notably, high densities of sequence reads from Xanthomonas translucens and Pseudomonas syringae were associated with some populations of Selaginella but not Lycopodium. Since these bacteria are already in use as biocontrol agents elsewhere, further investigation into their potential as biocontrol of Selaginella in New Zealand is suggested.

Effects of arbuscular mycorrhizal symbiosis on growth, nutrient and metal uptake by maize seedlings (Zea mays L.) grown in soils spiked with Lanthanum and Cadmium.

Multiple contaminants can affect plant-microbial remediation processes because of their interactive effects on environmental behaviour, bioavailability and plant growth. Recent studies have suggested that arbuscular mycorrhizal fungi (AMF) can facilitate the revegetation of soils co-contaminated with rare earth elements (REEs) and heavy metals. However, little is known regarding the role of AMF in the interaction of REEs and heavy metals. A pot experiment was conducted to evaluate the effects of Claroideoglomus etunicatum on the biomass, nutrient uptake, metal uptake and translocation of maize grown in soils spiked with Lanthanum (La) and Cadmium (Cd). The results indicated that individual and combined applications of La (100 mg kg-1) and Cd (5 mg kg-1) significantly decreased root colonization rates by 22.0%-35.0%. With AMF inoculation, dual-metal treatment significantly increased maize biomass by 26.2% compared to single-metal treatment. Dual-metal treatment significantly increased N, P and K uptake by 20.1%-76.8% compared to single-metal treatment. Dual-metal treatment significantly decreased shoot La concentration by 52.9% compared to single La treatment, whereas AM symbiosis caused a greater decrease of 87.8%. Dual-metal treatment significantly increased shoot and root Cd concentrations by 65.5% and 58.7% compared to single Cd treatment and the La translocation rate by 142.0% compared to single La treatment, whereas no difference was observed between their corresponding treatments with AMF inoculation. Furthermore, AMF had differential effects on the interaction of La and Cd on metal uptake and translocation under the background concentrations of soil metals. Taken together, these results indicated that AMF significantly affected the interaction between La and Cd, depending on metal types and concentrations in soils. These findings promote a further understanding of the contributions of AMF to the phytoremediation of co-contaminated soil.

Development and characterization of EST-SSR markers in Stipa breviflora (Poaceae).

PREMISE OF THE STUDY: Stipa breviflora (Poaceae) is one of the dominant species of the desert steppe in the eastern Eurasian grasslands. Simple sequence repeat (SSR) markers were developed for use in genetic diversity studies of this species. METHODS AND RESULTS: A total of 1954 potentially polymorphic loci were obtained by comparing transcriptome data of eight different S. breviflora individuals. We selected 81 loci to verify polymorphism and 63 loci amplified, of which 21 loci exhibited polymorphism. The number of alleles per locus varied from two to 24, the observed heterozygosity ranged from 0.083 to 0.958, and the expected heterozygosity ranged from 0.396 to 0.738. CONCLUSIONS: These newly identified SSR loci can be used for population genetic and landscape genetic studies of S. breviflora. In addition, 14 loci also amplified in six related Stipa species (S. grandis, S. krylovii, S. bungeana, S. aliena, S. gobica, and S. purpurea).

Gene transcript profiles in the desert plant Nitraria tangutorum during fruit development and ripening.

Nitraria tangutorum Bobr., a valuable wild shrub distributed in Northwest China, produces edible and medicinal berries. However, little is known about the molecular mechanisms of its fruit development and ripening. We performed de novo transcriptome sequencing of N. tangutorum fruit using the Illumina HiSeq™ 2000 sequencing platform. More than 62.94 million reads were obtained and assembled into 69,306 unigenes (average length, 587 bp). These unigenes were annotated by querying against five databases (Nr, Swiss-Prot, GO, COG, and KEGG); 42,929 and 26,809 unigenes were found in the Nr and Swiss-Prot databases, respectively. In ortholog analyses, 33,363 unigenes were assigned with one or more GO terms, 15,537 hits were aligned to 25 COG classes, and 24,592 unigenes were classified into 128 KEGG pathways. Digital gene expression analyses were conducted on N. tangutorum fruit at the green (S1), yellow (S2), and red (S3) developmental stages. In total, 8240, 5985, and 4994 differentially expressed genes (DEGs) were detected for S1 vs. S2, S1 vs. S3, and S2 vs. S3, respectively. Cluster analyses showed that a large proportion of DEGs related to plant hormones and transcription factors (TFs) showed high expression in S1, down-regulated expression in S2, and up-regulated expression in S3. We analyzed the expression patterns of 23 genes encoding 12 putative enzymes involved in flavonoid biosynthesis. The expression profiles of 10 DEGs involved in flavonoid biosynthesis were validated by Q-PCR analysis. The assembled and annotated transcriptome sequences and gene expression profile analyses provide valuable genetic resources for research on N. tangutorum.

Isolation and characterization of a Δ1-pyrroline-5-carboxylate synthetase (NtP5CS) from Nitraria tangutorum Bobr. and functional comparison with its Arabidopsis homologue.

Several functional and regulatory proteins play important roles in controlling plant stress tolerance. Proline (Pro) is one of the most accumulated osmolytes correlated with tolerance to stresses. Δ(1)-Pyrroline-5-carboxylate synthetase (P5CS) is a rate-limiting enzyme in Pro biosynthesis. In the present study, we isolated the cDNA for a P5CS gene (NtP5CS) from the halophyte Nitraria tangutorum. Phylogenetic analysis and subcellular localization analysis of NtP5CS-GFP protein in onion cells showed that NtP5CS was a new P5CS gene and was involved in Pro synthesis in N. tangutorum. Expression of the NtP5CS gene was induced by salt stress, dehydration, and high and low temperatures. Escherichia coli overexpressing AtP5CS or NtP5CS exhibited better growth in all treatments, including high salinity, high alkalinity, dehydration, osmotic, heat and cold stresses. Additionally, NtP5CS recombinant E. coli cells grew better than did AtP5CS recombinant cells in response to abiotic stresses. Our data demonstrate that the P5CS from a halophytic species functions more efficiently than its homologue from a glycophytic species in improving the stress tolerance of E. coli.

Identification of Salt-Stress-Induced Genes from the RNA-Seq Data of Reaumuria trigyna Using Differential-Display Reverse Transcription PCR.

Next generation sequencing (NGS) technologies have been used to generate huge amounts of sequencing data from many organisms. However, the correct choice of candidate genes and prevention of false-positive results computed from digital gene expression (DGE) of RNA-seq data are vital when using these genetic resources. We indirectly identified 18 salt-stress-induced Reaumuria trigyna transcripts from the transcriptome sequencing data using differential-display reverse transcription PCR (DDRT-PCR) combined with local BLAST searches. Highly consistent with the DGE results, the quantitative real-time PCR expression patterns of these transcripts showed strong upregulation by salt stress, suggesting that these genes may play important roles in R. trigyna's survival under high-salt environments. The method presented here successfully identified responsive genes from the massive amount of RNA-seq data. Thus, we suggest that DDRT-PCR could be employed to mine NGS data in a wide range of applications in transcriptomic studies. In addition, the genes identified in the present study are promising candidates for further elucidation of the salt tolerance mechanisms in R. trigyna.

Transcriptomic profiling of the salt-stress response in the wild recretohalophyte Reaumuria trigyna.

BACKGROUND: Reaumuria trigyna is an endangered small shrub endemic to desert regions in Inner Mongolia. This dicotyledonous recretohalophyte has unique morphological characteristics that allow it to tolerate the stress imposed by semi-desert saline soil. However, it is impossible to explore the mechanisms underlying this tolerance without detailed genomic information. Fortunately, newly developed high-throughput sequencing technologies are powerful tools for de novo sequencing to gain such information for this species. RESULTS: Two sequencing libraries prepared from control (C21) and NaCl-treated samples (T43) were sequenced using short reads sequencing technology (Illumina) to investigate changes in the R. trigyna transcriptome in response to salt stress. Among 65340 unigenes, 35495 (52.27%) were annotated with gene descriptions, conserved domains, gene ontology terms, and metabolic pathways with a cut-off E-value of 10-5. These included 44 Gene Ontology (GO) terms, 119 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways, and 25 Clusters of Orthologous Groups families. By comparing the transcriptomes from control and NaCl-treated plants, 5032 genes showed significantly differences in transcript abundance under salt stress (false discovery rate ≤ 0.001 and |log2Ratio| ≥ 1). These genes were significantly enriched in 29 KEGG pathways and 26 GO terms. The transcription profiles indicated that genes related to ion transport and the reactive oxygen species scavenging system were relevant to the morphological and physiological characteristics of this species. The expression patterns of 30 randomly selected genes resulted from quantitative real-time PCR were basically consistent with their transcript abundance changes identified by RNA-seq. CONCLUSIONS: The present study identified potential genes involved in salt tolerance of R. trigyna. The globally sequenced genes covered a considerable proportion of the R. trigyna transcriptome. These data represent a genetic resource for the discovery of genes related to salt tolerance in this species, and may be a useful source of reference sequences for closely related taxa. These results can also further our understanding of salt tolerance in other halophytes surviving under sodic stress.