Unique regulatory network of dragon fruit simultaneously mitigates the effect of vanadium pollutant and environmental factors.

Under changing climatic conditions, plants are simultaneously facing conflicting stresses in nature. Plants can sense different stresses, induce systematic ROS signals, and regulate transcriptomic, hormonal, and stomatal responses. We performed transcriptome analysis to reveal the integrative stress response regulatory mechanism underlying heavy metal stress alone or in combination with heat and drought conditions in pitaya (dragon fruit). A total of 70 genes were identified from 31,130 transcripts with conserved differential expression. Furthermore, weighted gene co-expression network analysis (WGCNA) identified trait-associated modules. By integrating information from three modules and protein-protein interaction (PPI) networks, we identified 10 interconnected genes associated with the multifaceted defense mechanism employed by pitaya against co-occurring stresses. To further confirm the reliability of the results, we performed a comparative analysis of 350 genes identified by three trait modules and 70 conserved genes exhibiting their dynamic expression under all treatments. Differential expression pattern of genes and comparative analysis, have proven instrumental in identifying ten putative structural genes. These ten genes were annotated as PLAT/LH2, CAT, MLP, HSP, PB1, PLA, NAC, HMA, and CER1 transcription factors involved in antioxidant activity, defense response, MAPK signaling, detoxification of metals and regulating the crosstalk between the complex pathways. Predictive analysis of putative candidate genes, potentially governing single, double, and multifactorial stress response, by several signaling systems and molecular patterns. These findings represent a valuable resource for pitaya breeding programs, offering the potential to develop resilient "super pitaya" plants.

Chloroplast Genomes Evolution and Phylogenetic Relationships of Caragana species.

Caragana sensu lato (s.l.) includes approximately 100 species that are mainly distributed in arid and semi-arid regions. Caragana species are ecologically valuable for their roles in windbreaking and sand fixation. However, the taxonomy and phylogenetic relationships of the genus Caragana are still unclear. In this study, we sequenced and assembled the chloroplast genomes of representative species of Caragana and reconstructed robust phylogenetic relationships at the section level. The Caragana chloroplast genome has lost the inverted repeat region and wascategorized in the inverted repeat loss clade (IRLC). The chloroplast genomes of the eight species ranged from 128,458 bp to 135,401 bp and contained 110 unique genes. All the Caragana chloroplast genomes have a highly conserved structure and gene order. The number of long repeats and simple sequence repeats (SSRs) showed significant variation among the eight species, indicating heterogeneous evolution in Caragana. Selective pressure analysis of the genes revealed that most of the protein-coding genes evolved under purifying selection. The phylogenetic analyses indicated that each section forms a clade, except the section Spinosae, which was divided into two clades. This study elucidated the evolution of the chloroplast genome within the widely distributed genus Caragana. The detailed information obtained from this study can serve as a valuable resource for understanding the molecular dynamics and phylogenetic relationships within Caragana.

Chloroplast genomes of Caragana tibetica and Caragana turkestanica: structures and comparative analysis.

BACKGROUND: The genus Caragana encompasses multiple plant species that possess medicinal and ecological value. However, some species of Caragana are quite similar in morphology, so identifying species in this genus based on their morphological characteristics is considerably complex. In our research, illumina paired-end sequencing was employed to investigate the genetic organization and structure of Caragana tibetica and Caragana turkestanica, including the previously published chloroplast genome sequence of 7 Caragana plants. RESULTS: The lengths of C. tibetica and C. turkestanica chloroplast genomes were 128,433 bp and 129,453 bp, respectively. The absence of inverted repeat sequences in these two species categorizes them under the inverted repeat loss clade (IRLC). They encode 110 and 111 genes (4 /4 rRNA genes, 30 /31tRNA genes, and 76 /76 protein-coding genes), respectively. Comparison of the chloroplast genomes of C. tibetica and C. turkestanica with 7 other Caragana species revealed a high overall sequence similarity. However, some divergence was observed between certain intergenic regions (matK-rbcL, psbD-psbM, atpA-psbI, and etc.). Nucleotide diversity (π) analysis revealed the detection of five highly likely variable regions, namely rps2-atpI, accD-psaI-ycf4, cemA-petA, psbN-psbH and rpoA-rps11. Phylogenetic analysis revealed that C. tibetica's sister species is Caragana jubata, whereas C. turkestanica's closest relative is Caragana arborescens. CONCLUSIONS: The present study provides worthwhile information about the chloroplast genomes of C. tibetica and C. turkestanica, which aids in the identification and classification of Caragana species.

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.

Multiple configurations of the plastid and mitochondrial genomes of Caragana spinosa.

MAIN CONCLUSION: In this study, we assembled the complete plastome and mitogenome of Caragana spinosa and explored the multiple configurations of the organelle genomes. Caragana spinosa belongs to the Papilionoidea subfamily and has significant pharmaceutical value. To explore the possible interaction between the organelle genomes, we assembled and analyzed the plastome and mitogenome of C. spinosa using the Illumina and Nanopore DNA sequencing data. The plastome of C. spinosa was 129,995 bp belonging to the inverted repeat lacking clade (IRLC), which contained 77 protein-coding genes, 29 tRNA genes, and four rRNA genes. The mitogenome was 378,373 bp long and encoded 54 unique genes, including 33 protein-coding, three ribosomal RNA (rRNA), and 18 transfer RNA (tRNA) genes. In addition to the single circular conformation, alternative conformations mediated by one and four repetitive sequences in the plastome and mitogenome were identified and validated, respectively. The inverted repeat (PDR12, the 12th dispersed repeat sequence in C. spinosa plastome) of plastome mediating recombinant was conserved in the genus Caragana. Furthermore, we identified 14 homologous fragments by comparing the sequences of mitogenome and plastome, including eight complete tRNA genes. A phylogenetic analysis of protein-coding genes extracted from the plastid and mitochondrial genomes revealed congruent topologies. Analyses of sequence divergence found one intergenic region, trnN-GUU-ycf1, exhibiting a high degree of variation, which can be used to develop novel molecular markers to distinguish the nine Caragana species accurately. This plastome and mitogenome of C. spinosa could provide critical information for the molecular breeding of C. spinosa and be used as a reference genome for other species of Caragana. In this study, we assembled the complete plastome and mitogenome of Caragana spinosa and explored the multiple configurations of the organelle genomes.

Genome-Wide Analysis of WRKY Transcription Factors Involved in Abiotic Stress and ABA Response in Caragana korshinskii.

The WRKY transcription factor family plays a vital role in plant development and environmental response. However, the information of WRKY genes at the genome-wide level is rarely reported in Caragana korshinskii. In this study, we identified and renamed 86 CkWRKY genes, which were further classified into three groups through phylogenetic analysis. Most of these WRKY genes were clustered and distributed on eight chromosomes. Multiple sequence alignment revealed that the conserved domain (WRKYGQK) of the CkWRKYs was basically consistent, but there were also six variation types (WRKYGKK, GRKYGQK, WRMYGQK, WRKYGHK, WKKYEEK and RRKYGQK) that appeared. The motif composition of the CkWRKYs was quite conservative in each group. In general, the number of WRKY genes gradually increased from lower to higher plant species in the evolutionary analysis of 28 species, with some exceptions. Transcriptomics data and RT-qPCR analysis showed that the CkWRKYs in different groups were involved in abiotic stresses and ABA response. Our results provided a basis for the functional characterization of the CkWRKYs involved in stress resistance in C. korshinskii.

Years of sand fixation with Caragana korshinskii drive the enrichment of its rhizosphere functional microbes by accumulating soil N.

C. korshinskii is one of the most widely-planted sand-fixing legumes in northwest China and exploring its rhizosphere microbiome is of great ecological importance. However, the effect of long-term sand fixation on the composition, diversity, and underlying functions of microbes in the C. korshinskii rhizosphere in dryland ecosystems remain unclear. Here, we performed high-throughput sequencing using a 16S rRNA (absolute quantification) and bacterial functional annotation of prokaryotic taxa (FAPROTAX) analysis and an ITS (relative quantification) and fungal functional guild (FUNGuild) analysis to investigate the C. korshinskii rhizosphere microbiome and metabolic functional groups at different sand-fixing ages (six years, CK6; twelve years, CK12; and eighteen years, CK18) and determined the physicochemical properties of the rhizosphere soil. Results showed that the key bacterial taxa of the rhizosphere were significantly more abundant in CK18 than in CK12 and CK6 at the phylum-class-genus level, and that fungal Glomeromycota was also significantly more abundant in the CK18 rhizosphere compared to CK12 and CK6. Among these bacterial taxa, the enrichment effect of key, functional, genus-level species of bacteria was the most obvious, including Rhizobium, Ensifer, Neorhizobium, Mesorhizobium, Streptomyces, Sphingomonas, and Flavobacterium, which are N-fixing and/or phosphate-solubilizing groups. The significant improvement seen in the physicochemical properties of the CK18 rhizosphere soil, including the higher total nitrogen (TN), available nitrogen (AN), pH, electrical conductivity (EC), higher N:P ratio, and lower C:N ratio, all demonstrated the relationship between the rhizosphere microbes and soil carbon (C) and nitrogen (N) cycling. A redundancy analysis (RDA) of different taxonomic levels indicated a close positive relationship between rhizosphere microbes and AN. In addition, the functional groups of the C. korshinskii rhizosphere bacteria were closely related to soil AN and were mainly composed of chemoheterotrophy and aerobic chemoheterotrophy. A Spearman correlation analysis revealed that these functional groups were mainly identified from bacterial Actinobacteria, Proteobacteria, Verrucomicrobia, Bacteroidetes, and fungal Glomeromycota. Our study provides evidence that the rhizosphere microbes of C. korshinskii are closely related to the accumulation of N in the restoration of desert ecosystems, and that the ecological functional processes they are involved in mainly involve C and N cycles, which play an important role in desertification reversal.

The chloroplasts genomic analyses of four specific Caragana species.

BACKGROUND: Many species of the genus Caragana have been used as wind prevention and sand fixation plants. They are also important traditional Chinese medicine, and ethnic medicine resource plant. Thus, chloroplast genomes (cp-genome) of some of these important species must be studied. METHODS: In this study, we analyzed the chloroplast genomes of C. jubata, C. erinacea, C. opulens, and C. bicolor, including their structure, repeat sequences, mutation sites, and phylogeny. RESULTS: The size of the chloroplast genomes was between 127,862 and 132,780 bp, and such genomes contained 112 genes (30 tRNA, 4 rRNA, and 78 protein-coding genes), 43 of which were photosynthesis-related genes. The total guanine + cytosine (G+C) content of four Caragana species was between 34.49% and 35.15%. The four Caragana species all lacked inverted repeats and can be classified as inverted repeat-lacking clade (IRLC). Of the anticipated genes of the four chloroplast genomes, introns were discovered in 17 genes, most of which were inserted by one intron. A total of 50 interspersed repeated sequences (IRSs) were found among them, 58, 29, 61, and 74 simple sequences repeats were found in C. jubata, C. bicolor, C. opulens, and C. erinacea, respectively. Analyses of sequence divergence showed that some intergenic regions (between trnK-UUU and rbcl; trnF-GAA and ndhJ; trnL-CAA and trnT-UGU; rpoB and trnC-GCA; petA and psbL; psbE and pebL; and sequences of rpoC, ycf1, and ycf2) exhibited a high degree of variations. A phylogenetic tree of eight Caragana species and another 10 legume species was reconstructed using full sequences of the chloroplast genome. CONCLUSIONS: (1) Chloroplast genomes can be used for the identification and classification of Caragana species. (2) The four Caragana species have highly similar cpDNA G+C content. (3) IRS analysis of the chloroplast genomes showed that these four species, similar to the chloroplast genome of most legumes, lost IRLC regions. (4) Comparative cp-genomic analysis suggested that the cp genome structure of the Caragana genus was well conserved in highly variable regions, which can be used to exploit markers for the identification of Caragana species and further phylogenetic study. (5) Results of phylogenetic analyses were in accordance with the current taxonomic status of Caragana. The phylogenetic relationship of Caragana species was partially consistent with elevation and geographical distribution.

CkREV Enhances the Drought Resistance of Caragana korshinskii through Regulating the Expression of Auxin Synthetase Gene CkYUC5.

As a common abiotic stress, drought severely impairs the growth, development, and even survival of plants. Here we report a transcription factor, Caragana korshinskii REVOLUTA(CkREV), which can bidirectionally regulate the expression of the critical enzyme gene CkYUC5 in auxin synthesis according to external environment changes, so as to control the biosynthesis of auxin and further enhance the drought resistance of plants. Quantitative analysis reveals that the expression level of both CkYUC5 and AtYUC5 is down-regulated after C. korshinskii and Arabidopsis thaliana are exposed to drought. Functional verification of CkREV reveals that CkREV up-regulates the expression of AtYUC5 in transgenic A. thaliana under common conditions, while down-regulating it under drought conditions. Meanwhile, the expression of CkYUC5 is also down-regulated in C. korshinskii leaves instantaneously overexpressing CkREV. We apply a dual-luciferase reporter system to discover that CkREV can bind to the promoter of CkYUC5 to regulate its expression, which is further proved by EMSA and Y1H esxperiments. Functional verification of CkREV in C. korshinskii and transgenic A. thaliana shows that CkREV can regulate the expression of CkYUC5 and AtYUC5 in a contrary way, maintaining the equilibrium of plants between growth and drought resisting. CkREV can positively regulate the expression of CkYUC5 to promote auxin synthesis in favor of growth under normal development. However, CkREV can also respond to external signals and negatively regulate the expression of CkYUC5, which inhibits auxin synthesis in order to reduce growth rate, lower water demands, and eventually improve the drought resistance of plants.

Overexpression of γ-glutamylcysteine synthetase gene from Caragana korshinskii decreases stomatal density and enhances drought tolerance.

BACKGROUND: Gamma-glutamylcysteine synthetase (γ-ECS) is a rate-limiting enzyme in glutathione biosynthesis and plays a key role in plant stress responses. In this study, the endogenous expression of the Caragana korshinskii γ-ECS (Ckγ-ECS) gene was induced by PEG 6000-mediated drought stress in the leaves of C. korshinskii. and the Ckγ-ECS overexpressing transgenic Arabidopsis thaliana plants was constructed using the C. korshinskii. isolated γ-ECS. RESULTS: Compared with the wildtype, the Ckγ-ECS overexpressing plants enhanced the γ-ECS activity, reduced the stomatal density and aperture sizes; they also had higher relative water content, lower water loss, and lower malondialdehyde content. At the same time, the mRNA expression of stomatal development-related gene EPF1 was increased and FAMA and STOMAGEN were decreased. Besides, the expression of auxin-relative signaling genes AXR3 and ARF5 were upregulated. CONCLUSIONS: These changes suggest that transgenic Arabidopsis improved drought tolerance, and Ckγ-ECS may act as a negative regulator in stomatal development by regulating the mRNA expression of EPF1 and STOMAGEN through auxin signaling.

Transcriptome-based identification and expression profiling of AP2/ERF members in Caragana intermedia and functional analysis of CiDREB3.

BACKGROUND: The AP2/ERF transcription factor family plays important roles in regulation of plant growth and development as well as the response of plants to stress. However, there are currently few studies focusing on the function of the AP2/ERF-type transcription factors in Caragana intermedia Kuang et H. C. Fu. Here, the expression pattern of AP2/ERF transcription factors family in different tissues and under four stress treatments were evaluated, and the function of CiDREB3 was examined. METHODS AND RESULTS: In this study, the genes encoding the AP2/ERF family of transcription factors were screened from the C. intermedia drought transcriptome database and subjected to bioinformatic analysis using the online tool and software. The expression pattern of the members of AP2/ERF transcription factors in C. intermedia were detected via quantitative real-time PCR (qRT-PCR). The function of CiDREB3 on growth, development and drought tolerance was evaluated by transgenic Arabidopsis. As a result, 22 sequences with complete ORFs were obtained and all sequences were divided into 13 sub-groups. Most of the AP2/ERF transcription factors exhibited tissue-specific expression and were induced by cold, heat, NaCl and mannitol treatments. Furthermore, heterologous expression of CiDREB3 altered the morphology of the transgenic Arabidopsis thaliana L. Heynh and improved its drought tolerance during seedlings development. CONCLUSIONS: Taken together, the results of the present study helped to better understand the function of the AP2/ERF family transcription factors in response to multiple abiotic stresses and uncovered the role of CiDREB3 in affecting the morphology and abiotic stress tolerance of Arabidopsis.

Enhanced tolerance to drought stress resulting from Caragana korshinskii CkWRKY33 in transgenic Arabidopsis thaliana.

BACKGROUND: It is well known that WRKY transcription factors play important roles in plant growth and development, defense regulation and stress responses. RESULTS: In this study, a WRKY transcription factor, WRKY33, was cloned from Caragana korshinskii. A sequence structure analysis showed that it belonged to the Group-I type. Subcellular localization experiments in tobacco epidermal cells showed the presence of CkWRKY33 in the nucleus. Additionally, CkWRKY33 was overexpressed in Arabidopsis thaliana. A phenotypic investigation revealed that compared with wild-type plants, CkWRKY33-overexpressing transgenic plants had higher survival rates, as well as relative soluble sugar, proline and peroxidase contents, but lower malondialdehyde contents, following a drought stress treatment. CONCLUSIONS: This suggested that the overexpression of CkWRKY33 led to an enhanced drought-stress tolerance in transgenic A. thaliana. Thus, CkWRKY33 may act as a positive regulator involved in the drought-stress responses in Caragana korshinskii.

Functional characterization of an unobtrusive protein, CkMT4, in re-establishing desiccation tolerance in germinating seeds.

Desiccation tolerance (DT) is gradually lost during seed germination, while it can be re-established by pre-treatment with polyethylene glycol (PEG) and/or abscisic acid (ABA). Increasing knowledge is available on several stress-related proteins in DT re-establishment in herb seeds, but limited information exists on novel proteins in wood seeds. This study aimed to investigate the role of metallothionein CkMT4, a protein species with the highest fold increase in abundance in Caragana korshinskii seeds on PEG treatment. The fluctuation in mRNA levels of CkMT4 during seed development was consistent with the changes in DT, and the expression of CkMT4 could be up-regulated by ABA. Besides metal-binding capacity, CkMT4 might supply Cu2+/Zn2+ to superoxide dismutase (SOD) under high redox potential provided by PEG treatment for excess reactive oxygen species (ROS) scavenging. The overexpression of CkMT4 in yeast results in enhanced oxidation resistance. Experimentally, this study demonstrated the overexpression of CkMT4 in Arabidopsis seeds benefited the re-establishment of DT and enhanced the activity of SOD. On the whole, these findings suggested that CkMT4 facilitated the re-establishment of DT in C. korshinskii seeds mainly through diminishing excess ROS, which put the mechanism underlying the re-establishment of DT in xerophytic wood seeds into a new perspective.

Comprehensive transcriptome profiling of Caragana microphylla in response to salt condition using de novo assembly.

OBJECTIVE: To investigate the response of Caragana microphylla in salt condition, transcriptome analysis, differentially expressed genes (DEGs) comparison with Arabidopsis thaliana, and the chlorophyll content analysis were performed. RESULTS: Gene Ontology (GO) term, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses of DEGs indicated that salt condition affected photosynthesis and chlorophyll in C. microphylla. The DEGs compared with salt responsive genes of A. thaliana indicated that C. microphylla's responses to salt differed greatly from those of the model plant and that the results also indicated up-regulated genes related to photosynthesis and chlorophyll in C. microphylla. Moreover, we confirmed that salt-treated C. microphylla increased chlorophyll content, and the genes of protoporphyrin IX downstream in chlorophyll biosynthesis were induced in the heatmap analysis. CONCLUSIONS: These results showed a similar pattern to some halophytes plants with increased chlorophyll at a certain salt concentration, and we assumed that C. microphylla also has a mechanism to adapt or tolerate moderate salt conditions.

Identification of drought response genes by digital gene expression (DGE) analysis in Caragana korshinskii Kom.

Caragana korshinskii Kom. is a legume shrub that is widely distributed across desert habitats with gravely, sandy, and saline soils in Asia and Africa. C. korshinskii has highly developed roots and a strong tolerance to abiotic stress. At present, there are few genetic studies of C. korshinskii because of the limited availability of genomic resources. To understand the comprehensive mechanisms that are associated with drought tolerance, we used RNA-seq to survey the differentially expressed genes (DEGs) in comparisons of drought-treated and control plants. After analysing the sequencing results, we found 440 differentially expressed genes existing in drought-treated and control plants. Among the DEGs, 39 unigenes showed up-regulated expression after drought treatment, while 401 unigenes were down-regulated. We used the KEGG database to annotate these drought-induced genes; 126 unigenes were identified by KEGG pathway annotation, and approximately 28% of the unigenes with known function fell into categories related to fatty acid metabolism, starch, sucrose metabolism, and nitrogen metabolism, suggesting that these pathways or processes may be involved in the drought response. Finally, we confirmed that one gene has a potential function in drought tolerance. Our study is the first to provide transcriptomic resources for Caragana korshinskii and to determine its digital gene expression profile under conditions of drought stress using the assembled transcriptomic data for reference. These data provide a valuable resource for genetic and genomic studies of desert plants under abiotic stress conditions.

Development of Agrobacterium-mediated transient expression system in Caragana intermedia and characterization of CiDREB1C in stress response.

BACKGROUND: The Agrobacterium-mediated transient transformation is a versatile and indispensable way of rapid analyzing gene function in plants. Despite this transient expression system has been successfully applied in a number of plant species, it is poorly developed in Caragana intermedia. RESULTS: In this study, we established an Agrobacterium-mediated transient expression system in C. intermedia leaves and optimized the effect of different Agrobacterial strains, several surfactants and the concentration of Silwet L-77, which would affect transient expression efficiency. Among the 5 Agrobacterial strains examined, GV3101 produced the highest GUS expression level. Besides, higher level of transient expression was observed in plants infiltrated with Silwet L-77 than with Triton X-100 or Tween-20. Silwet L-77 at a concentration of 0.001% greatly improved the level of GUS transient expression. Real-time PCR showed that expression of CiDREB1C was highly up-regulated in transiently expressed plants and reached the highest level at the 2nd day after infiltration. Based on this optimized transient transformation method, we characterized CiDREB1C function in response to drought, salt and ABA treatment. The results showed that transiently expressed CiDREB1C in C. intermedia leaves could enhance the survival rate and chlorophyll content, and reduce the lodging rate compared with the control seedlings under drought, salt and ABA treatments. Furthermore, the rate of leaf shedding of CiDREB1C transient expression seedlings was lower than that of the control under ABA treatment. CONCLUSIONS: The optimized transient expression condition in C. intermedia leaves were infiltrated with Agrobacterial strains GV3101 plus Silwet L-77 at a concentration of 0.001% added into the infiltration medium. Transiently expressed CiDREB1C enhanced drought, salt and ABA stress tolerance, indicated that it was a suitable and effective tool to determine gene function involved in abiotic stress response in C. intermedia.

Sequencing, Characterization, and Comparative Analyses of the Plastome of Caragana rosea var. rosea.

To exploit the drought-resistant Caragana species, we performed a comparative study of the plastomes from four species: Caragana rosea, C. microphylla, C. kozlowii, and C. Korshinskii. The complete plastome sequence of the C. rosea was obtained using the next generation DNA sequencing technology. The genome is a circular structure of 133,122 bases and it lacks inverted repeat. It contains 111 unique genes, including 76 protein-coding, 30 tRNA, and four rRNA genes. Repeat analyses obtained 239, 244, 258, and 246 simple sequence repeats in C. rosea, C. microphylla, C. kozlowii, and C. korshinskii, respectively. Analyses of sequence divergence found two intergenic regions: trnI-CAU-ycf2 and trnN-GUU-ycf1, exhibiting a high degree of variations. Phylogenetic analyses showed that the four Caragana species belong to a monophyletic clade. Analyses of Ka/Ks ratios revealed that five genes: rpl16, rpl20, rps11, rps7, and ycf1 and several sites having undergone strong positive selection in the Caragana branch. The results lay the foundation for the development of molecular markers and the understanding of the evolutionary process for drought-resistant characteristics.

What drives the shift between sexual and clonal reproduction of Caragana stenophylla along a climatic aridity gradient?

BACKGROUND: The reasons that clonal plants shift between sexual and clonal reproduction have persisted as a knowledge gap in ecological literature. We hypothesized that clonal plants' shifts between sexual and clonal reproduction in different environments are driven by the relative costs of sexual and clonal reproduction. Moreover, we hypothesized plants prioritize sexual reproduction over clonal reproduction. To test these hypotheses, we determined the costs of sexual and clonal reproduction, and proportions of sexual and clonal reproduction of Caragana stenophylla along a climatic aridity gradient (semi-arid, arid, very arid and intensively arid zones) in the Inner Mongolia Steppe using several complementary field experiments. RESULTS: The cost of sexual reproduction increased while the cost of clonal reproduction decreased as climatic drought stress increased from the semi-arid to the intensively arid zones. The changes in the costs of these reproductive modes drove a shift in the reproductive mode of C. stenophylla from more sexual reproduction in the semi-arid zone to more clonal propagation in the intensively arid zone. However, because of the evolutionary advantages of sexual reproduction, sexual reproduction still held priority over clonal production in C. stenophylla, with the priority of sexual reproduction gradually increasing from the semi-arid to the intensively arid zones. CONCLUSIONS: Our study suggested that sexual reproduction has relatively high priority in propagation of C. stenophylla. However, if the costs of sexual reproduction are too high, C. stenophylla likely chooses clonal reproduction, and the ratio between sexual and clonal reproduction could be mediated by reproductive cost. These reproductive strategies reflect optimal resource utilization, and allow the persistence of both reproductive modes across stressful conditions depending on their evolutionary advantages.

Identification of the WRKY gene family and functional analysis of two genes in Caragana intermedia.

BACKGROUND: WRKY transcription factors, one of the largest families of transcriptional regulators in plants, play important roles in plant development and various stress responses. The WRKYs of Caragana intermedia are still not well characterized, although many WRKYs have been identified in various plant species. RESULTS: We identified 53 CiWRKY genes from C. intermedia transcriptome data, 28 of which exhibited complete open reading frames (ORFs). These CiWRKYs were divided into three groups via phylogenetic analysis according to their WRKY domains and zinc finger motifs. Conserved domain analysis showed that the CiWRKY proteins contain a highly conserved WRKYGQK motif and two variant motifs (WRKYGKK and WKKYEEK). The subcellular localization of CiWRKY26 and CiWRKY28-1 indicated that these two proteins localized exclusively to nuclei, supporting their role as transcription factors. The expression patterns of the 28 CiWRKYs with complete ORFs were examined through quantitative real-time PCR (qRT-PCR) in various tissues and under different abiotic stresses (drought, cold, salt, high-pH and abscisic acid (ABA)). The results showed that each CiWRKY responded to at least one stress treatment. Furthermore, overexpression of CiWRKY75-1 and CiWRKY40-4 in Arabidopsis thaliana suppressed the drought stress tolerance of the plants and delayed leaf senescence, respectively. CONCLUSIONS: Fifty-three CiWRKY genes from the C. intermedia transcriptome were identified and divided into three groups via phylogenetic analysis. The expression patterns of the 28 CiWRKYs under different abiotic stresses suggested that each CiWRKY responded to at least one stress treatment. Overexpression of CiWRKY75-1 and CiWRKY40-4 suppressed the drought stress tolerance of Arabidopsis and delayed leaf senescence, respectively. These results provide a basis for the molecular mechanism through which CiWRKYs mediate stress tolerance.

Population genetic structure is shaped by historical, geographic, and environmental factors in the leguminous shrub Caragana microphylla on the Inner Mongolia Plateau of China.

BACKGROUND: Understanding how landscape factors, including suites of geographic and environmental variables, and both historical and contemporary ecological and evolutionary processes shape the distribution of genetic diversity is a primary goal of landscape and conservation genetics and may be particularly consequential for species involved in ecological restoration. In this study, we examine the factors that shape the distribution of genetic variation in a leguminous shrub (Caragana microphylla) important for restoration efforts on the Mongolian Plateau in China. This region houses several major bioclimatic gradients, and C. microphylla is an important restoration species because it stabilizes soils and prevents advancing desertification on the Inner Mongolia Plateau caused by ongoing climate change. RESULTS: We assembled an expansive genomic dataset, consisting of 22 microsatellite loci, four cpDNA regions, and 5788 genome-wide SNPs from ten populations of C. microphylla. We then applied ecological niche modelling and linear and non-linear regression techniques to investigate the historical and contemporary forces that explain patterns of genetic diversity and population structure in C. microphylla on the Inner Mongolia Plateau. We found strong evidence that both geographic and environmental heterogeneity contribute to genetic differentiation and that the spatial distribution of genetic diversity in C. microphylla appears to result partly from the presence of a glacial refugium at the southwestern edge of its current range. CONCLUSIONS: These results suggest that geographic, environmental, and historical factors have all contributed to spatial genetic variation in this ecologically important species. These results should guide restoration plans to sustain genetic diversity during plant translocations.