Genome diversity and highland-adaptative variation in Tibet barley landrace population of China.

Barley landraces accumulated variation in adapting to extreme highland environments during long-term domestication in Tibet, but little is known about their population structure and genomic selection traces. In this study, tGBS (tunable genotyping by sequencing) sequencing, molecular marker and phenotypic analyses were conducted on 1,308 highland and 58 inland barley landraces in China. The accessions were divided into six sub-populations and clearly distinguished most six-rowed, naked barley accessions (Qingke in Tibet) from inland barley. Genome-wide differentiation was observed in all five sub-populations of Qingke and inland barley accessions. High genetic differentiation in the pericentric regions of chromosomes 2H and 3H contributed to formation of five types of Qingke. Ten haplotypes of the pericentric regions of 2H, 3H, 6H and 7H were further identified as associated with ecological diversification of these sub-populations. There was genetic exchange between eastern and western Qingke but they shared the same progenitor. The identification of 20 inland barley types indicated multiple origins of Qingke in Tibet. The distribution of the five types of Qingke corresponded to specific environments. Two predominant highland-adaptative variations were identified for low temperature tolerance and grain color. Our results provide new insights into the origin, genome differentiation, population structure and highland adaptation in highland barley which will benefit both germplasm enhancement and breeding of naked barley.

Comparative metabolomics analysis of the response to cold stress of resistant and susceptible Tibetan hulless barley (Hordeum distichon).

Plants cultivated on the Qinghai-Tibetan Plateau grow in an extremely cold environment and thus are exposed to cold stress. To assess the metabolic processes during cold exposure of Tibetan hulless barley (Hordeum distichon L.), metabolic analyses were conducted on one tolerant (XiLa) and one sensitive (ZangQing) cultivar exposed to six temperatures (24 °C, 12 °C, 5 °C, 0 °C, -5 °C, -8 °C) for 24 h. In total, 770 metabolites were identified, including amino acids and derivatives, carbohydrates, flavonoids, lipids, nucleotides and derivatives, and phenolamides. In principal component analysis, the samples were clearly grouped according to the cultivar, suggesting that the two cultivars have differential responses to cold stress. In cold-grown plants, eight metabolites, including monoacylglycerol (MAG, 18:2), MAG (18:3), deoxyadenosine, 6-methylmercaptopurine, and coniferin, were significantly altered in XiLa, but not in ZangQing when compared to the levels in control plants, and thus, these compounds can be considered as potential biomarkers of exposure to cold stress in hulless barley. Furthermore, differentially altered metabolites between seedlings exposed to -8 °C and those maintained at 24 °C were significantly enriched in glutathione metabolism. The findings of this study will be useful for the development of cultivars with cold stress tolerance.

Comparative proteomics analysis of Tibetan hull-less barley under osmotic stress via data-independent acquisition mass spectrometry.

BACKGROUND: Tibetan hull-less barley (Hordeum vulgare L. var. nudum) is one of the primary crops cultivated in the mountains of Tibet and encounters low temperature, high salinity, and drought. Specifically, drought is one of the major abiotic stresses that affect and limit Tibetan barley growth. Osmotic stress is often simultaneously accompanied by drought conditions. Thus, to improve crop yield, it is critical to explore the molecular mechanism governing the responses of hull-less barley to osmotic/drought stress conditions. FINDINGS: In this study, we used quantitative proteomics by data-independent acquisition mass spectrometry to investigate protein abundance changes in tolerant (XL) and sensitive (DQ) cultivars. A total of 6,921 proteins were identified and quantified in all samples. Two distinct strategies based on pairwise and time-course comparisons were utilized in the comprehensive analysis of differentially abundant proteins. Further functional analysis of differentially abundant proteins revealed that some hormone metabolism-associated and phytohormone abscisic acid-induced genes are primarily affected by osmotic stress. Enhanced regulation of reactive oxygen species (may promote the tolerance of hull-less barley under osmotic stress. Moreover, we found that some regulators, such as GRF, PR10, MAPK, and AMPK, were centrally positioned in the gene regulatory network, suggesting that they may have a dominant role in the osmotic stress response of Tibetan barley. CONCLUSIONS: Our findings highlight a subset of proteins and processes that are involved in the alleviation of osmotic stress. In addition, this study provides a large-scale and multidimensional proteomic data resource for the further investigation and improvement of osmotic/drought stress tolerance in hull-less barley or other plant species.

Metabolite profiling in two contrasting Tibetan hulless barley cultivars revealed the core salt-responsive metabolome and key salt-tolerance biomarkers.

Salinity stress represents one of the most harmful abiotic stresses for agricultural productivity. Tibetan hulless barley is an important economic crop widely grown in highly stressful conditions in the Qinghai-Tibet Plateau and is often challenged by salinity stress. To investigate the temporal metabolic responses to salinity stress in hulless barley, we performed a widely targeted metabolomic analysis of 72 leaf samples from two contrasting cultivars. We identified 642 compounds 57 % of which were affected by salt stress in the two cultivars, principally amino acids and derivatives, organic acids, nucleotides, and derivatives and flavonoids. A total of 13 stress-related metabolites including piperidine, L-tryptophan, L-glutamic acid, L-saccharopine, L-phenylalanine, 6-methylcoumarin, cinnamic acid, inosine 5'-monophosphate, aminomalonic acid, 6-aminocaproic acid, putrescine, tyramine and abscisic acid (ABA) represent the core metabolome responsive to salinity stress in hulless barley regardless of the tolerance level. In particular, we found that the ABA signalling pathway is essential to salt stress response in hulless barley. The high tolerance of the cultivar 0119 is due to a metabolic reprogramming at key stress times. During the early salt stress stages (0-24 h), 0119 tended to save energy through reduced glycolysis, nucleotide metabolism and amino acid synthesis, while increased antioxidant compounds such as flavonoids. Under prolonged stress (48-72 h), 0119 significantly enhanced energy production and amino acid synthesis. In addition, some important compatible solutes were strongly accumulated. By comparing the two cultivars, nine salt-tolerance biomarkers, mostly unreported salt-tolerance compounds in plants, were uncovered. Our study indicated that the salt tolerant hulless barley cultivar invokes a tolerance strategy which is conserved in other plant species. Overall, we provide for the first time some extensive metabolic data and some important salt-tolerance biomarkers which may assist in efforts to improve hulless barley tolerance to salinity stress.

Insights into the metabolic responses of two contrasting Tibetan hulless barley genotypes under low nitrogen stress.

Nitrogen (N) is an essential macronutrient for plants. However, excessive use of N fertilizer for cultivation is an environmental hazard. A good adaption to N deficiency is known in the Tibetan hulless barley. Therefore, it is of interest to complete the metabolic analysis on LSZQK which is a low nitrogen (low-N) sensitive genotype and Z0284 that is tolerant to low-N. We identified and quantified 750 diverse metabolites in this analysis. The two genotypes show differences in their basal metabolome under normal N condition. Polyphenols and lipids related metabolites were significantly enriched in Z0284 having a basal role prior to exposure to low-N stress. Analysis of the differentially accumulated metabolites (DAM) induced by low-N explain the genotype-specific responses. Fourteen DAMs showed similar patterns of change between low-N and control conditions in both genotypes. This could be the core low-N responsive metabolites regardless of the tolerance level in hulless barley. We also identified 4 DAMs (serotonin, MAG (18:4) isomer 2, tricin 7-O-feruloylhexoside and gluconic acid) shared by both genotypes displaying opposite patterns of regulation under low-N conditions and may play important roles in low-N tolerance. This report provides a theoretical basis for further understanding of the molecular mechanisms of low-N stress tolerance in hulless barley.

Gene coexpression network analysis combined with metabonomics reveals the resistance responses to powdery mildew in Tibetan hulless barley.

Powdery mildew is a fungal disease that represents a ubiquitous threat to crop plants. Transcriptomic and metabolomic analyses were used to identify molecular and physiological changes in Tibetan hulless barley in response to powdery mildew. There were 3418 genes and 405 metabolites differentially expressed between the complete resistance cultivar G7 and the sensitive cultivar Z13. Weighted gene coexpression network analysis was carried out, and the differentially expressed genes were enriched in five and four major network modules in G7 and Z13, respectively. Further analyses showed that phytohormones, photosynthesis, phenylpropanoid biosynthesis, and flavonoid biosynthesis pathways were altered during Qingke-Blumeria graminis (DC.) f.sp. hordei (Bgh) interaction. Comparative analyses showed a correspondence between gene expression and metabolite profiles, and the activated defenses resulted in changes of metabolites involved in plant defense response, such as phytohormones, lipids, flavone and flavonoids, phenolamides, and phenylpropanoids. This study enabled the identification of Bgh responsive genes and provided new insights into the dynamic physiological changes that occur in Qingke during response to powdery mildew. These findings greatly improve our understanding of the mechanisms of induced defense response in Qingke and will provide new clues for the development of resistant Tibetan hulless barley varieties.

Time-Course Comparative Metabolite Profiling under Osmotic Stress in Tolerant and Sensitive Tibetan Hulless Barley.

Tibetan hulless barley is widely grown in the extreme environmental conditions of the Qinghai-Tibet Plateau which is characterized by cold, high salinity, and drought. Osmotic stress always occurs simultaneously with drought and its tolerance is a vital part of drought tolerance. The diversity of metabolites leading to osmotic stress tolerance was characterized using widely-targeted metabolomics in tolerant (XL) and sensitive (D) accessions submitted to polyethylene glycol. XL regulated a more diverse set of metabolites than D, which may promote the establishment of a robust system to cope with the stress in XL. Compounds belonging to the group of flavonoids, amino acids, and glycerophospholipids constitute the core metabolome responsive to the stress, despite the tolerance levels. Moreover, 8 h appeared to be a critical time point for stress endurance involving a high accumulation of key metabolites from the class of nucleotide and its derivative which provide the ultimate energy source for the synthesis of functional carbohydrates, lipids, peptides, and secondary metabolites in XL. This intrinsic metabolic adjustment helped XL to efficiently alleviate the stress at the later stages. A total of 22 diverse compounds were constantly and exclusively regulated in XL, representing novel stress tolerance biomarkers which may help improving stress tolerance, especially drought, in hulless barley.

Study on content determination of lobetyolin and gallic acid in Eighteen Flavors Dangshen Pill from different factories / 重庆医学

Objective To develop a HPLC method for determining the contents of lobetyolin and gallic acid in Eighteen Flavors Dangshen Pill(EFDSP) produced by different factories.Methods The HPLC analysis was performed on a VP-DOS C18 column (4.6 mm×150 mm,5 μm).The mobile phase was acetonitrile and 0.4% glacial acetic acid(21∶79) in the determination of lobetyolin content.The detection wavelength was 267 nm and the flow velocity was 1 mL/min.the column temperature was25 ℃ and the sample size was10 μL.The mobile phase was methanol and 0.4% glacial acetic acid(1∶99) in the determination of gallic acid content.The detection wavelength was 280 nm.The column temperature was 25 ℃ and the sample size was 10μL.Results The contents of lobetyolin and gallic acid in EFDSP were 1.0835mg·g-1and 15.334 0 mg/g for Qinghai Gela Dandong Tibetan Pharmaceutical Factory;0.628 9 mg/g and 15.159 5 mg/g for Changdu Tibet Medicine Factory;0.306 5 mg/g and 8.762 7 mg/g for Tibetan Hospital of Tibet.Conclusion This method has the advantages of good reproducibility,good accuracy,simple and fast operation.The contents of lobetyolin and gallic acid in EFDSP produced by different manufacturers are significantly different.The gallic acid content has greater difference.It provides the reference for quality control of EFDSP