Differential Regulations of Antioxidant Metabolism and Cold-Responsive Genes in Three Bermudagrass Genotypes under Chilling and Freezing Stress.

As a typical warm-season grass, bermudagrass growth and turf quality begin to decrease when the environmental temperature drops below 20 °C. The current study investigated the differential responses of three bermudagrass genotypes to chilling stress (8/4 °C) for 15 days and then freezing stress (2/-2 °C) for 2 days. The three genotypes exhibited significant variation in chilling and freezing tolerance, and Chuannong-3, common bermudagrass 001, and Tifdwarf were ranked as cold-tolerant, -intermediate, and -sensitive genotypes based on evaluations of chlorophyll content, the photochemical efficiency of photosystem II, oxidative damage, and cell membrane stability, respectively. Chuannong-3 achieved better tolerance through enhancing the antioxidant defense system to stabilize cell membrane and reactive oxygen species homeostasis after being subjected to chilling and freezing stresses. Chuannong-3 also downregulated the ethylene signaling pathway by improving CdCTR1 expression and suppressing the transcript levels of CdEIN3-1 and CdEIN3-2; however, it upregulated the hydrogen sulfide signaling pathway via an increase in CdISCS expression under cold stress. In addition, the molecular basis of cold tolerance could be associated with the mediation of key genes in the heat shock pathway (CdHSFA-2b, CdHSBP-1, CdHSP22, and CdHSP40) and the CdOSMOTIN in Chuannong-3 because the accumulation of stress-defensive proteins, including heat shock proteins and osmotin, plays a positive role in osmoprotection, osmotic adjustment, or the repair of denatured proteins as molecular chaperones under cold stress. The current findings give an insight into the physiological and molecular mechanisms of cold tolerance in the new cultivar Chuannong-3, which provides valuable information for turfgrass breeders and practitioners.

Evaluation of zinc tolerance and accumulation in eight cultivars of bermudagrass (Cynodon spp.): implications for zinc phytoremediation.

Zinc (Zn) is a vital element for plant growth and development, however, excessive Zn is toxic to plants. Common bermudagrass (Cynodon dactylon (L.) Pers.) and hybrid bermudagrass (C. dactylon (L.) Pers. × C. transvaalensis Burtt-Davy) are widely used turfgrass species with strong tolerance to diverse abiotic stresses, including excessive Zn2+ stress. However, the variation of zinc tolerance and accumulation in different bermudagrass cultivars remain unclear. In this study, we systematically analyzed the growth performance, physiological index and ion concentration in eight commercial cultivars of common and hybrid bermudagrass under different concentration of Zn2+ treatments using pot experiments. The results indicated that four cultivars of common bermudagrass could tolerate 20 mM Zn2+, whereas four cultivars of hybrid bermudagrass could only tolerate 10 mM Zn2+. Among the four common bermudagrass cultivars, cultivar Guanzhong and Common showed stronger Zn tolerance and accumulation abilities than other two cultivars. Further analyses of the expression of selected Zn homeostasis-related genes indicated that bermudagrass cultivars with stronger tolerance to excessive Zn have at least one expression-elevated gene involved in Zn homeostasis. These results not only expanded our understanding of Zn tolerance and accumulation in bermudagrass but also facilitated the application of commercial bermudagrass cultivars in phytoremediation of Zn pollution.

Overexpression of CdtCIPK21 from triploid bermudagrass reduces salt and drought tolerance but increases chilling tolerance in transgenic rice.

Calcineurin B-like-interacting protein kinase (CIPK) is a serine/threonine kinase, which transmits the Ca2+ signal sensed by CBL proteins. A CdtCIPK21 showing highly identical to OsCIPK21 in rice was isolated from triploid bermudagrass (Cynodon dactylon × Cynodon transvaalensis). CdtCIPK21 transcript could be detected in roots, rhizomes, stems, stolons, and leaves, with highest level in roots. It was induced by salinity, dehydration and chilling, but reduced by ABA treatment. Transgenic rice plants overexpressing CdtCIPK21 had decreased salt and drought tolerance as well as ABA sensitivity but increased chilling tolerance. Lower SOD and CAT activities was observed in transgenic lines under salinity and drought stress conditions, but higher levels under chilling stress. Similarly, lower levels of proline concentration and P5CS1 and P5CS2 transcripts were maintained in transgenic lines under salinity and drought stresses, and higher levels were maintained under chilling. In addition, transgenic lines had lower transcript levels of ABA-independent genes (OsDREB1A, OsDREB1B, and OsDREB2A) and ABA responsive genes (OsLEA3, OsLIP9, and OsRAB16A) under salinity and drought but higher levels under chilling compared with WT. The results suggest that CdtCIPK21 regulates salt and drought tolerance negatively and chilling tolerance positively, which are associated with the altered ABA sensitivity, antioxidants, proline accumulation and expression of ABA-dependent and ABA-independent stress responsive genes.

Leaf Cuticular Waxes of Bermudagrass Response to Environment-Driven Adaptations of Climate Effect Inferred from Latitude and Longitude Gradient in China.

Bermudagrass (Cynodon dactylon) is a widely used warm season lawn grass. Cuticular wax covering the surface of plant leaves plays an important role in helping plants resist biotic and abiotic stresses. We analyzed the changes of cuticle wax in 25 bermudagrass populations from different longitude and latitude gradients, in order to verify how environmental conditions affect the structure and chemical composition of cuticle wax. Five wax components were identified, including alkanes, esters, alkenes, aldehydes and primary alcohols. The wax characteristics were divided into two principal components, explaining 58.2 % and 66.7 % of the total variability in latitude and longitude, even some populations had a certain correlation with each other. Pearson correlation analysis further showed that the total wax coverage, wax component content and antioxidant enzyme activity of bermudagrass populations on the latitudinal gradient had different responses to environmental factors. Finally, nineteen key genes involved in wax biosynthesis, redox and photosynthesis were identified and verified by RT-qPCR. The results showed that the responses of bermudagrass in different populations to climate change were quite different, which was of great significance for the evolution of bermudagrass populations.

miRNAs and Their Target Genes Play a Critical Role in Response to Heat Stress in Cynodon dactylon (L.) Pers.

Annual global temperature is increasing rapidly. Therefore, in the near future, plants will be exposed to severe heat stress. However, the potential of microRNAs-mediated molecular mechanism for modulating the expression of their target genes is unclear. To investigate the changes of miRNAs in thermo-tolerant plants, in this study, we first investigated the impact of four high temperature regimes including 35/30 °C, 40/35 °C, 45/40 °C, and 50/45 °C in a day/night cycle for 21 days on the physiological traits (total chlorophyll, relative water content and electrolyte leakage and total soluble protein), antioxidant enzymes activities (superoxide dismutase, ascorbic peroxidase, catalase and peroxidase), and osmolytes (total soluble carbohydrates and starch) in two bermudagrass accessions named Malayer and Gorgan. The results showed that more chlorophyll and the relative water content, lower ion leakage, more efficient protein and carbon metabolism and activation of defense proteins (such as antioxidant enzymes) in Gorgan accession, led to better maintained plant growth and activity during heat stress. In the next stage, to investigate the role of miRNAs and their target genes in response to heat stress in a thermo-tolerant plant, the impact of severe heat stress (45/40 °C) was evaluated on the expression of three miRNAs (miRNA159a, miRNA160a and miRNA164f) and their target genes (GAMYB, ARF17 and NAC1, respectively). All measurements were performed in leaves and roots simultaneously. Heat stress significantly induced the expression of three miRNAs in leaves of two accession, while having different effects on the expression of these miRNAs in roots. The results showed that a decrease in the expression of the transcription factor ARF17, no change in the expression of the transcription factor NAC1, and an increase in the expression of the transcription factor GAMYB in leaf and root tissues of Gorgan accession led to improved heat tolerance in it. These results also showed that the effect of miRNAs on the modulating expression of target mRNAs in leaves and roots is different under heat stress, and miRNAs and mRNAs show spatiotemporal expression. Therefore, the simultaneous analysis of miRNAs and mRNAs expressions in shoot and roots is needed to comprehensively understand miRNAs regulatory function under heat stress.

The extended day length promotes earlier flowering of bermudagrass.

Day length is a very critical environmental factor affecting plant growth and development. The extension of light application time has been shown to promote flowering in the long-day plant and to shorten breeding time in some crops. However, previous research on the regulation of bermudagrass flowering by light application time is scarce. Therefore, this study investigated the effect of day length on the growth and flowering of bermudagrass by prolonging the light application time in a controlled greenhouse. Three different light application times were set up in the experiment: 22/2 h (22 hours light/2 hours dark), 18/6 h (18 hours light/6 hours dark), 14/10 h (14 hours light/10 hours dark). Results showed that extending the light application time not only promoted the growth of bermudagrass (plant height, fresh weight, dry weight) but also its nutrient uptake (nitrogen (N) and phosphorous (P) content). In addition, daily light integrals were different when flowering under different light application times. Most importantly, under the 22/2 h condition, flowering time was successfully reduced to 44 days for common bermudagrass (Cynodon dactylon [L.] pers) genotype A12359 and 36 days for African bermudagrass (Cynodon transvaalensis Burtt-Davy) genotype ABD11. This study demonstrated a successful method of bermudagrass flowering earlier than usual time by manipulating light application time which may provide useful insights for bermudagrass breeding.

A Novel Beta-Glucosidase Gene for Plant Type Was Identified by Genome-Wide Association Study and Gene Co-Expression Analysis in Widespread Bermudagrass.

Bermudagrass (Cynodon spp.) is one of the most widely distributed warm-season grasses globally. The growth habits and plant type of bermudagrass are strongly associated with the applied purpose of the landscape, livestock, and eco-remediation. Therefore, persistent efforts are made to investigate the genetic basis of plant type and growth habits of bermudagrass. Here, we dissect the genetic diversity of 91 wild bermudagrass resources by genome-wide association studies (GWAS) combined with weighted gene co-expression analysis (WGCNA). This work is based on the RNA-seq data and the genome of African bermudagrass (Cynodon transvaalensis Burtt Davy). Sixteen reliable single-nucleotide polymorphisms (SNPs) in transcribed regions were identified to be associated with the plant height and IAA content in diverse bermudagrass by GWAS. The integration of the results from WGCNA indicates that beta-glucosidase 31 (CdBGLU31) is a candidate gene underlying a G/A SNP signal. Furthermore, both qRT-PCR and correlation coefficient analyses indicate that CdBGLU31 might play a comprehensive role in plant height and IAA biosynthesis and signal. In addition, we observe lower plant height in Arabidopsis bglu11 mutants (homologs of CdBGLU31). It uncovers the breeding selection history of different plant types from diverse bermudagrass and provides new insights into the molecular function of CdBGLU31 both in plant types and in IAA biosynthetic pathways.

Proteome-wide analyses reveal diverse functions of protein acetylation and succinylation modifications in fast growing stolons of bermudagrass (Cynodon dactylon L.).

BACKGROUND: Bermudagrass (Cynodon dactylon L.) is an important warm-season turfgrass species with well-developed stolons, which lay the foundation for the fast propagation of bermudagrass plants through asexual clonal growth. However, the growth and development of bermudagrass stolons are still poorly understood at the molecular level. RESULTS: In this study, we comprehensively analyzed the acetylation and succinylation modifications of proteins in fast-growing stolons of the bermudagrass cultivar Yangjiang. A total of 4657 lysine acetylation sites on 1914 proteins and 226 lysine succinylation sites on 128 proteins were successfully identified using liquid chromatography coupled to tandem mass spectrometry, respectively. Furthermore, 78 proteins and 81 lysine sites were found to be both acetylated and succinylated. Functional enrichment analysis revealed that acetylated proteins regulate diverse reactions of carbohydrate metabolism and protein turnover, whereas succinylated proteins mainly regulate the citrate cycle. These results partly explained the different growth disturbances of bermudagrass stolons under treatment with sodium butyrate and sodium malonate, which interfere with protein acetylation and succinylation, respectively. Moreover, 140 acetylated proteins and 42 succinylated proteins were further characterized having similarly modified orthologs in other grass species. Site-specific mutations combined with enzymatic activity assays indicated that the conserved acetylation of catalase and succinylation of malate dehydrogenase both inhibited their activities, further implying important regulatory roles of the two modifications. CONCLUSION: In summary, our study implied that lysine acetylation and succinylation of proteins possibly play important regulatory roles in the fast growth of bermudagrass stolons. The results not only provide new insights into clonal growth of bermudagrass but also offer a rich resource for functional analyses of protein lysine acetylation and succinylation in plants.

Comparative analysis reveals chromosome number reductions in the evolution of African bermudagrass (Cynodon transvaalensis Burtt-Davy).

African bermudagrass (Cynodon transvaalensis Burtt-Davy) (2n = 2x = 18) belongs to the genus Cynodon, tribe Cynodonteae, subfamily Chloridoideae in the grass family Poaceae. The species is frequently crossed with common bermudagrass (Cynodon dactylon Pers.) in developing high-quality hybrid turf cultivars. Molecular resources for C. transvaalensis are scarce; thus, its genomic evolution is unknown. Recently, a linkage map consisting of 1278 markers provided a powerful tool for African bermudagrass genomic research. The objective of this study was to investigate chromosome number reduction events that resulted in the nine haploid chromosomes in this species. Tag sequences of mapped single nucleotide polymorphism markers in C. transvaalensis were compared against genome sequences of Oropetium thomaeum (L.f.) Trin. (2n = 2x = 20), a genomic model in the Cynodonteae tribe. The comparative genomic analyses revealed broad collinearity between the genomes of these two species. The analyses further revealed that two major interchromosomal rearrangements of the paleochromosome ρ12 (ρ1-ρ12-ρ1 and ρ6-ρ12-ρ6) resulted in nine chromosomes in the genome of C. transvaalensis. The findings provide novel information regarding the formation of the initial diploid species in the Cynodon genus.

Genetic manipulation of bermudagrass photosynthetic biosynthesis using Agrobacterium-mediated transformation.

Bermudagrass is one of the most extensively used warm-season grasses. It is widely used in landscaping, stadium construction and soil remediation due to its excellent regeneration, trampling and stress tolerances. However, studies on its regulatory mechanism and variety improvement by genetic engineering are still at a standstill, owing to its genetic variability and intrinsic limits linked with some resistance to Agrobacterium infection. In this study, we established a higher efficient Agrobacterium-mediated transformation via screening for vital embryogenic callus and improving infection efficiency. The superior callus was light yellow, hard granular and compact, determined with a differentiation rate of more than 95%. The optimized infestation courses by gentle shaking, vacuuming and sonicating were used. The infested calluses were co-cultured for 3 days, followed by desiccation treatments for 1 day to get higher infection efficiency. Then the CdHEMA1 gene, essential for chlorophyll biosynthesis, was cloned and transferred into bermudagrass to validate the aforementioned optimization procedures integrally. Molecular-level analyses indicated that the CdHEMA1 gene had successfully integrated and was greatly increased in transgenic seedlings. Results of the photosynthetic capacity assessment showed that CdHEMA1 overexpression may considerably enhance the contents of photosynthetic pigments, OJIP curve and reaction center density (RC/CSo) to absorb (ABS/CSo, ABS/CSM) and capture (TRo/CSo) more light energy, hence improve the performance indices PIABS and PICS compared to the wild type. The successful completion of this project would provide a solid platform for further gene function study and molecular breeding of bermudagrass.

Physiological integration between Bermudagrass ramets improves overall salt resistance under heterogeneous salt stress.

Connected ramets of colonal plants often suffer from different environmental conditions such as light, nutrient, and stress. Colonal Bermudagrass (Cynodon dactylon [L.] Pers.) can form interconnected ramets and this connection facilitates the tolerance to abiotic stress, which is a kind of physiological integration. However, how bermudagrass responds to heterogeneously distributed salt stress needs to be further elucidated. Here, we demonstrated that severance of stolons aggravated the damage of salt-stressed ramets, displaying higher relative electrolytic leakage (EL), lower content of chlorophyll, higher accumulation of Na+ , and serious oxidative damages. This finding implied the positive effects of the physiological integration of bermudagrass on salt tolerance. The unstressed ramets connected with the stressed one were mildly injured, implying the supporting and sacrifice function of the unstressed ramets. Physiological integration did not mediate the translocation of Na+ among ramets, but induced a higher expression of salt overly sensitive (SOS) genes in the stressed ramets, consequently reducing the accumulation of Na+ in leaves and roots. In addition, physiological integration upregulated the genes expression and enzymes activity of catalase (CAT) and peroxidase (POD) in both stressed and unstressed ramets. This granted a stronger antioxidant ability of the whole clonal plants under salt stress. Enhanced Na+ transfer and increased reactive oxygen species (ROS) scavenging are mechanisms that likely contribute to the physiological integration leading to the salt tolerance of bermudagrass.

CdWRKY2-mediated sucrose biosynthesis and CBF-signalling pathways coordinately contribute to cold tolerance in bermudagrass.

Bermudagrass (Cynodon dactylon) is one of the most widely cultivated warm-season turfgrass species around the world. Cold stress has been a key environmental factor that adversely affects the growth, development, and geographical distribution of bermudagrass; however, the underlying mechanism of bermudagrass responsive to cold stress remains largely unexplored. Here, we identified a cold-induced WRKY transcription factor CdWRKY2 from bermudagrass and demonstrated its function in cold stress response. Overexpression of CdWRKY2 enhanced cold tolerance in transgenic Arabidopsis and bermudagrass hairy roots, while knocking down CdWRKY2 expression via virus-induced gene silencing increased cold susceptibility. RNA sequencing showed that overexpression of CdWRKY2 in Arabidopsis activated the expression of genes involved in sucrose synthesis and metabolism, including sucrose synthase 1 (AtSUS1) and sucrose phosphate synthase 2F (AtSPS2F). CdSPS1, the homology gene of AtSPS2F in bermudagrass, was subsequently proven to be the direct target of CdWRKY2 by yeast one-hybrid, electrophoretic mobility shift assay, and transient expression analysis. As expected, overexpression of CdSPS1 conferred cold tolerance in transgenic Arabidopsis plants, whereas silencing CdSPS1 expression enhanced cold sensitivity in bermudagrass. Besides, CdCBF1 whose expression was dramatically up-regulated in CdWRKY2-overexpressing bermudagrass hairy roots but down-regulated in CdWRKY2-silencing bermudagrass both under normal and cold stress conditions was confirmed as another target of CdWRKY2. Collectively, this study reveals that CdWRKY2 is a positive regulator in cold stress by targeting CdSPS1 and CdCBF1 promoters and activating their expression to coordinately mediate sucrose biosynthesis and CBF-signalling pathway, which provides valuable information for breeding cold-resistant bermudagrass through gene manipulation.

Integration of the metabolome and transcriptome reveals the mechanism of resistance to low nitrogen supply in wild bermudagrass (Cynodon dactylon (L.) Pers.) roots.

BACKGROUND: Nitrogen (N) is an essential macronutrient that significantly affects turf quality. Commercial cultivars of bermudagrass (Cynodon dactylon (L.) Pers.) require large amounts of nitrogenous fertilizer. Wild bermudagrass germplasm from natural habitats with poor nutrition and diverse N distributions is an important source for low-N-tolerant cultivated bermudagrass breeding. However, the mechanisms underlying the differences in N utilization among wild germplasm resources of bermudagrass are not clear. RESULTS: To clarify the low N tolerance mechanism in wild bermudagrass germplasm, the growth, physiology, metabolome and transcriptome of two wild accessions, C291 (low-N-tolerant) and C716 (low-N-sensitive), were investigated. The results showed that root growth was less inhibited in low-N-tolerant C291 than in low-N-sensitive C716 under low N conditions; the root dry weight, soluble protein content and free amino acid content of C291 did not differ from those of the control, while those of C716 were significantly decreased. Down-regulation of N acquisition, primary N assimilation and amino acid biosynthesis was less pronounced in C291 than in C716 under low N conditions; glycolysis and the tricarboxylic acid (TCA) cycle pathway were also down-regulated, accompanied by a decrease in the biosynthesis of amino acids; strikingly, processes such as translation, biosynthesis of the structural constituent of ribosome, and the expression of individual aminoacyl-tRNA synthetase genes, most of genes associated with ribosomes related to protein synthesis were all up-regulated in C291, but down-regulated in C716. CONCLUSIONS: Overall, low-N-tolerant wild bermudagrass tolerated low N nutrition by reducing N primary assimilation and amino acid biosynthesis, while promoting the root protein synthesis process and thereby maintaining root N status and normal growth.

Transcriptomic Analysis of the Photosynthetic, Respiration, and Aerenchyma Adaptation Strategies in Bermudagrass (Cynodon dactylon) under Different Submergence Stress.

Submergence impedes photosynthesis and respiration but facilitates aerenchyma formation in bermudagrass. Still, the regulatory genes underlying these physiological responses are unclear in the literature. To identify differentially expressed genes (DEGs) related to these physiological mechanisms, we studied the expression of DEGs in aboveground and underground tissues of bermudagrass after a 7 d treatment under control (CK), shallow submergence (SS), and deep submergence (DS). Results show that compared with CK, 12276 and 12559 DEGs were identified under SS and DS, respectively. Among them, the DEGs closely related to the metabolism of chlorophyll biosynthesis, light-harvesting, protein complex, and carbon fixation were down-regulated in SS and DS. Meanwhile, a large number of DEGs involved in starch and sucrose hydrolase activities, glycolysis/gluconeogenesis, tricarboxylic acid (TCA) cycle, and oxidative phosphorylation were down-regulated in aboveground tissues of bermudagrass in SS and DS. Whereas in underground tissues of bermudagrass these DEGs were all up-regulated under SS, only beta-fructofuranosidase and α-amylase related genes were up-regulated under DS. In addition, we found that DEGs associated with ethylene signaling, Ca2+-ROS signaling, and cell wall modification were also up-regulated during aerenchyma formation in underground tissues of bermudagrass under SS and DS. These results provide the basis for further exploration of the regulatory and functional genes related to the adaptability of bermudagrass to submergence.

Comprehensive transcriptional analysis reveals salt stress-regulated key pathways, hub genes and time-specific responsive gene categories in common bermudagrass (Cynodon dactylon (L.) Pers.) roots.

BACKGROUND: Despite its good salt-tolerance level, key genes and pathways involved with temporal salt response of common bermudagrass (Cynodon dactylon (L.) Pers.) have not been explored. Therefore, in this study, to understand the underlying regulatory mechanism following the different period of salt exposure, a comprehensive transcriptome analysis of the bermudagrass roots was conducted. RESULTS: The transcripts regulated after 1 h, 6 h, or 24 h of hydroponic exposure to 200 mM NaCl in the roots of bermudagrass were investigated. Dataset series analysis revealed 16 distinct temporal salt-responsive expression profiles. Enrichment analysis identified potentially important salt responsive genes belonging to specific categories, such as hormonal metabolism, secondary metabolism, misc., cell wall, transcription factors and genes encoded a series of transporters. Weighted gene co-expression network analysis (WGCNA) revealed that lavenderblush2 and brown4 modules were significantly positively correlated with the proline content and peroxidase activity and hub genes within these two modules were further determined. Besides, after 1 h of salt treatment, genes belonging to categories such as signalling receptor kinase, transcription factors, tetrapyrrole synthesis and lipid metabolism were immediately and exclusively up-enriched compared to the subsequent time points, which indicated fast-acting and immediate physiological responses. Genes involved in secondary metabolite biosynthesis such as simple phenols, glucosinolates, isoflavones and tocopherol biosynthesis were exclusively up-regulated after 24 h of salt treatment, suggesting a slightly slower reaction of metabolic adjustment. CONCLUSION: Here, we revealed salt-responsive genes belonging to categories that were commonly or differentially expressed in short-term salt stress, suggesting possible adaptive salt response mechanisms in roots. Also, the distinctive salt-response pathways and potential salt-tolerant hub genes investigated can provide useful future references to explore the molecular mechanisms of bermudagrass.

Genetic and QTL mapping in African bermudagrass.

Cynodon transvaalensis Burtt-Davy is frequently used to cross with C. dactylon Pers. in the creation of F1 hybrid cultivars that are some of the most widely used in the worldwide turf industry. However, molecular resource development in this species is limited. Accordingly, the objectives of this study were to construct a high-density genetic map, and to identify genomic regions associated with establishment rate. In this study, we constructed the first high-density linkage map for African bermudagrass using a genotyping by sequencing approach based on 109 S1 progenies. A total of 1,246 single nucleotide polymorphisms and 32 simple sequence repeat markers were integrated in the linkage map. The total length of nine linkage groups was 882.3 cM, with an average distance of 0.69 cM per interval. Four genomic regions were identified to be associated with sod establishment rate. The results provide important genetic resources towards understanding the genome as well as marker-assisted selection for improving the establishment rate in bermudagrass breeding.

Sequence-based genetic mapping of Cynodon dactylon Pers. reveals new insights into genome evolution in Poaceae.

Bermudagrass (Cynodon dactylon Pers.) is an important warm-season perennial used extensively for turf, forage, soil conservation and remediation worldwide. However, limited genomic information has hindered the application of molecular tools towards understanding genome evolution and in breeding new cultivars. We genotype a first-generation selfed population derived from the tetraploid (4x = 36) 'A12359' using genotyping-by-sequencing. A high-density genetic map of 18 linkage groups (LGs) is constructed with 3,544 markers. Comparative genomic analyses reveal that each of nine homeologous LG pairs of C. dactylon corresponds to one of the first nine chromosomes of Oropetium thomaeum. Two nested paleo-ancestor chromosome fusions (ρ6-ρ9-ρ6, ρ2-ρ10-ρ2) may have resulted in a 12-to-10 chromosome reduction. A segmental dissemination of the paleo-chromosome ρ12 (ρ1-ρ12-ρ1, ρ6-ρ12-ρ6) leads to the 10-to-9 chromosome reduction in C. dactylon genome. The genetic map will assist in an ongoing whole genome sequence assembly and facilitate marker-assisted selection (MAS) in developing new cultivars.

The ethylene responsive factor CdERF1 from bermudagrass (Cynodon dactylon) positively regulates cold tolerance.

Cold stress is one of the major environmental factors that limit growth and utilization of bermudagrass [Cynodon dactylon (L.) Pers], a prominent warm-season turfgrass. However, the molecular mechanism of cold response in bermudagrass remains largely unknown. In this study, we characterized a cold-responsive ERF (ethylene responsive factor) transcription factor, CdERF1, from bermudagrass. CdERF1 expression was induced by cold, drought and salinity stresses. The CdERF1 protein was nucleus-localized and encompassed transcriptional activation activity. Transgenic Arabidopsis plants overexpressing CdERF1 showed enhanced cold tolerance, whereas CdERF1-underexpressing bermudagrass plants via virus induced gene silencing (VIGS) method exhibited reduced cold resistance compared with control, respectively. Under cold stress, electrolyte leakage (EL), malondialdehyde (MDA), H2O2 and O2- contents were reduced, while the activities of SOD and POD were elevated in transgenic Arabidopsis. By contrast, these above physiological indicators in CdERF1-underexpressing bermudagrass exhibited the opposite trend. To further explore the possible molecular mechanism of bermudagrass cold stress response, the RNA-Seq analyses were performed. The result indicated that overexpression of CdERF1 activated a subset of stress-related genes in transgenic Arabidopsis, such as CBF2, pEARLI1 (lipid transfer protein), PER71 (peroxidase) and LTP (lipid transfer protein). Interestingly, under-expression of CdERF1 suppressed the transcription of many genes in CdERF1-underexpressing bermudagrass, also including pEARLI1 (lipid transfer protein) and PER70 (peroxidase). All these results revealed that CdERF1 positively regulates plant cold response probably by activating stress-related genes, PODs, CBF2 and LTPs. This study also suggests that CdERF1 may be an ideal candidate in the effort to improve cold tolerance of bermudagrass in the further molecular breeding.

Morphological variation in Cynodon dactylon (L.) Pers., and its relationship with the environment along a longitudinal gradient.

BACKGROUND: Geographical variation in morphological traits may reflect evolutionary patterns of morphological adaptability along environmental gradients. Comprehensive information on longitudinal patterns of morphological trait variation is very meaningful to explore morphological diversity and evolutionary trends in widespread bermudagrass. METHODS: To explore the spatial patterns of morphological traits, we investigated 10 morphological traits of bermudagrass and 10 soil nutrient indexes and collected local climate data for 13 different regions from 119°E to 105°E along the latitude 34°N. RESULTS: Considerable variations in morphological traits were observed at different longitudes, and the variations in most of the evaluated traits within populations were lower than those among populations. All of the 13 different longitudinal sites were divided into three groups based on morphological traits by cluster analysis. The major sources of diversity at the different longitudes were leaf length of the erect shoot, leaf width of the erect shoot, and the internode lengths of the erect shoot and stolon as determined by principal component analysis. Pearson correlation analysis also indicated that longitude was significantly and negatively correlated with these traits as well. Mean average rainfall was significantly correlated with leaf length of the erect shoot and the internode lengths of the erect shoot and stolon, while mean average temperature was only significantly correlated with internode length of the erect shoots. Available sulfur was significantly correlated with internode length of the erect shoot, plant height, and reproductive branch height, while the exchangeable Ca was significantly correlated with internode lengths of the erect shoot and stolon. Soil pH was significantly correlated with the internode length of the stolon. Longitude is an important factor that affects morphological trait variation in wild bermudagrass, and the leaves of the erect shoot and the internode length enlarged significantly with the collection sites moving from east to west. CONCLUSION: Different combinations and interactions of environmental factors (soil and climate) along a longitudinal gradient may have strong effects on one or more morphological traits of bermudagrass.

Protective functions of alternative splicing transcripts (CdDHN4-L and CdDHN4-S) of CdDHN4 from bermudagrass under multiple abiotic stresses.

Dehydrins (DHNs) play critical roles in plant adaptation to abiotic stresses. The objective of this study was to characterize DHNs in bermudagrass (Cynodon spp.). CdDHN4 gene was cloned from bermudagrass 'Tifway'. Two CdDHN4 transcripts were detected due to alternative splicing (the nonspliced CdDHN4-L and the spliced CdDHN4-S) and both the CdDHN4-S and CdDHN4-L proteins are YSK2-type DHNs, the Φ-segment is present in CdDHN4-L and absent in CdDHN4-S. Transgenic Arabidopsis thaliana expressing CdDHN4-L or CdDHN4-S exhibited improved tolerance to salt, osmotic, low temperature and drought stress compared to the wild type (WT). The two transgenic lines did not differ in salt or drought tolerance, while plants expressing CdDHN4-S grew better under osmotic stress than those expressing CdDHN4-L. Both transgenic lines exhibited reduced content of malondialdehyde (MDA) and reactive oxygen species (ROS); and higher antioxidant enzymatic activities than the wild type plants under salt or drought stress. CdDHN4-S exhibited a higher ROS-scavenging capacity than CdDHN4-L.