Validation of an Isocratic HPLC Method for Simultaneous Estimation of Major Phytosterols in Prunus spinosa L. extracts.

This study aimed to develop a rapid method for separation of stigmasterol, campesterol and ß-sitosterol in Prunus spinosa L. (sloe) fruit extracts by High Performance Liquid Chromatography system. Samples were prepared by Soxhlet extraction method and separated on a high strength silica C18 column using acetonitrile-methanol mobile phase and Photodiode Array Detector. The optimized method resulted in a linear calibration curve ranging from 1.7 ng mL-1 to 130 ng mL-1 for all three phytosterols. Analyses of internal and external phytosterol standards showed good linearity (R2 of 0.998 to 0.999); LOD and LOQ were determined to be 2.33×10-7-2.18×10-4 and 7.07×10-7-6.60×10-4 mg mL-1, respectively. Repeatability and reproducibility precision analyses showed acceptable values of RSD %. ß-sitosterol was the predominant phytosterol (51.53-81.03 % of total) among all samples. Method validation parameters indicated that this analytical method can be applied for accurate and precise determination of campesterol, stigmasterol and ß-sitosterol, in selected extracts.

Mandelonitrile lyase MDL2-mediated regulation of seed amygdalin and oil accumulation of Prunus Sibirica.

BACKGROUND: The Prunus sibirica seeds with rich oils has great utilization, but contain amygdalin that can be hydrolyzed to release toxic HCN. Thus, how to effectively reduce seed amygdalin content of P. sibirica is an interesting question. Mandelonitrile is known as one key intermediate of amygdalin metabolism, but which mandelonitrile lyase (MDL) family member essential for its dissociation destined to low amygdalin accumulation in P. sibirica seeds still remains enigmatic. An integration of our recent 454 RNA-seq data, amygdalin and mandelonitrile content detection, qRT-PCR analysis and function determination is described as a critical attempt to determine key MDL and to highlight its function in governing mandelonitrile catabolism with low amygdalin accumulation in Prunus sibirica seeds for better developing edible oil and biodiesel in China. RESULTS: To identify key MDL and to unravel its function in governing seed mandelonitrile catabolism with low amygdalin accumulation in P. sibirica. Global identification of mandelonitrile catabolism-associated MDLs, integrated with the across-accessions/developing stages association of accumulative amount of amygdalin and mandelonitrile with transcriptional level of MDLs was performed on P. sibirica seeds of 5 accessions to determine crucial MDL2 for seed mandelonitrile catabolism of P. sibirica. MDL2 gene was cloned from the seeds of P. sibirica, and yeast eukaryotic expression revealed an ability of MDL2 to specifically catalyze the dissociation of mandelonitrile with the ideal values of Km (0.22 mM) and Vmax (178.57 U/mg). A combination of overexpression and mutation was conducted in Arabidopsis. Overexpression of PsMDL2 decreased seed mandelonitrile content with an increase of oil accumulation, upregulated transcript of mandelonitrile metabolic enzymes and oil synthesis enzymes (involving FA biosynthesis and TAG assembly), but exhibited an opposite situation in mdl2 mutant, revealing a role of PsMDL2-mediated regulation in seed amygdalin and oil biosynthesis. The PsMDL2 gene has shown as key molecular target for bioengineering high seed oil production with low amygdalin in oilseed plants. CONCLUSIONS: This work presents the first integrated assay of genome-wide identification of mandelonitrile catabolism-related MDLs and the comparative association of transcriptional level of MDLs with accumulative amount of amygdalin and mandelonitrile in the seeds across different germplasms and developmental periods of P. sibirica to determine MDL2 for mandelonitrile dissociation, and an effective combination of PsMDL2 expression and mutation, oil and mandelonitrile content detection and qRT-PCR assay was performed to unravel a mechanism of PsMDL2 for controlling amygdalin and oil production in P. sibirica seeds. These findings could offer new bioengineering strategy for high oil production with low amygdalin in oil plants.

Mume Fructus reduces interleukin-1 beta-induced cartilage degradation via MAPK downregulation in rat articular chondrocytes.

Osteoarthritis is the most prevalent type of degenerative arthritis. It is characterized by persistent pain, joint dysfunction, and physical disability. Pain relief and inflammation control are prioritised during osteoarthritis treatment Mume Fructus (Omae), a fumigated product of the Prunus mume fruit, is used as a traditional medicine in several Asian countries. However, its therapeutic mechanism of action and effects on osteoarthritis and articular chondrocytes remain unknown. In this study, we analyzed the anti-osteoarthritis and articular regenerative effects of Mume Fructus extract on rat chondrocytes. Mume Fructus treatment reduced the interleukin-1ß-induced expression of matrix metalloproteinase 3, matrix metalloproteinase 13, and a disintegrin and metalloproteinase with thrombospondin type 1 motifs 5. Additionally, it enhanced collagen type II alpha 1 chain and aggrecan accumulation in rat chondrocytes. Furthermore, Mume Fructus treatment regulated the inflammatory cytokine levels, mitogen-activated protein kinase phosphorylation, and nuclear factor-kappa B activation. Overall, our results demonstrated that Mume Fructus inhibits osteoarthritis progression by inhibiting the nuclear factor-kappa B and mitogen-activated protein kinase pathways to reduce the levels of inflammatory cytokines and prevent cartilage degeneration. Therefore, Mume Fructus may be a potential therapeutic option for osteoarthritis.

Time-Separated Pulse Release-Activation of an Enzyme from Alginate-Polyethylenimine Hydrogels Using Electrochemically Generated Local pH Changes.

ß-Glucosidase (EC 3.2.1.21) from sweet almond was encapsulated into pH-responsive alginate-polyethylenimine (alginate-PEI) hydrogel. Then, electrochemically controlled cyclic local pH changes resulting from ascorbate oxidation (acidification) and oxygen reduction (basification) were used for the pulsatile release of the enzyme from the composite hydrogel. Activation of the enzyme was controlled by the very same pH changes used for ß-glucosidase release, separating these two processes in time. Importantly, the activity of the enzyme, which had not been released yet, was inhibited due to the buffering effect of PEI present in the gel. Thus, only a portion of the released enzyme was activated. Both enzymatic activity and release were monitored by confocal fluorescence microscopy and regular fluorescent spectroscopy. Namely, commercially available very little or nonfluorescent substrate 4-methylumbelliferyl-ß-d-glucopyranoside was hydrolyzed by ß-glucosidase to produce a highly fluorescent product 4-methylumbelliferone during the activation phase. At the same time, labeling of the enzyme with rhodamine B isothiocyanate was used for release observation. The proposed work represents an interesting smart release-activation system with potential applications in biomedical field.

A comprehensive review on pharmacognosy, phytochemistry and pharmacological activities of 8 potent species of southeast Asia.

Genus Prunus comprising around 430 species is a vast important genus of family Rosaceae, subfamily amygdalaoidae. Among all 430 species, around 19 important species are commonly found in Indian sub-continent due to their broad nutritional and economic importance. Some most common species of genus Prunus are Prunus amygdalus, Prunus persica, Prunus armeniaca, Prunus avium, Prunus cerasus, Prunus cerasoides, Prunus domestica, Prunus mahaleb, etc. A newly introduced species of Prunus i.e Prunus sunhangii is recently discovered which is morphologically very similar to Prunus cerasoides. Plants of Prunus species are short to medium-sized deciduous trees mainly found in the northern hemisphere. In India and its subcontinent, it extends from the Himalayas to Sikkim, Meghalaya, Bhutan, Myanmar etc. Different Prunus species have been extensively studied for their morphological, microscopic, pharmacological and phytoconstituents characteristics. Total phenolic content of Prunus species explains the presence of phenols in high quantity and pharmacological activity due to phenols. Phytochemical screening of species of genus Prunus shows the presence of wide phytoconstituents which contributes in their pharmacological significance and reveals the therapeutic potential and traditional medicinal significance of this genus. Genus Prunus showed a potent antioxidant activity analyzed by 1,1-diphenyl-2-picryl-hydrazyl radical assay. Plant species belonging to the genus Prunus is widely used traditionally for the treatment of various disorders. Some specific Prunus species possess potent anticancer, anti-inflammatory, hypoglycemic etc. activity which makes the genus more interesting for further research and findings. This review is an attempt to summarize the comprehensive study of Prunus.

Rapid determination of total flavonoid content, xanthine oxidase inhibitory activities, and antioxidant activity in Prunus mume by near-infrared spectroscopy.

Evaluating the quality of herbal medicine based on the content and activity of its main components is highly beneficial. Developing an eco-friendly determination method has significant application potential. In this study, we propose a new method to simultaneously predict the total flavonoid content (TFC), xanthine oxidase inhibitory (XO) activity, and antioxidant activity (AA) of Prunus mume using near-infrared spectroscopy (NIR). Using the sodium nitrite-aluminum nitrate-sodium hydroxide colorimetric method, uric acid colorimetric method, and 2,2-diphenyl-1-picrylhydrazyl radical (DPPH) free radical scavenging activity as reference methods, we analyzed TFC, XO, and AA in 90 P. mume samples collected from different locations in China. The solid samples were subjected to NIR. By employing spectral preprocessing and optimizing spectral bands, we established a rapid prediction model for TFC, XO, and AA using partial least squares regression (PLS). To improve the model's performance and eliminate irrelevant variables, competitive adaptive reweighted sampling (CARS) was used to calculate the pretreated full spectrum. Evaluation model indicators included the root mean square error of cross-validation (RMSECV) and determination coefficient (R2) values. The TFC, XO, and AA model, combining optimal spectral preprocessing and spectral bands, had RMSECV values of 0.139, 0.117, and 0.121, with RCV2 values exceeding 0.92. The root mean square error of prediction (RMSEP) for the TFC, XO, and AA model on the prediction set was 0.301, 0.213, and 0.149, with determination coefficient (RP2) values of 0.915, 0.933, and 0.926. The results showed a strong correlation between NIR with TFC, XO, and AA in P. mume. Therefore, the established model was effective, suitable for the rapid quantification of TFC, XO, and AA. The prediction method is simple and rapid, and can be extended to the study of medicinal plant content and activity.

Antigout effects and mechanisms of total flavonoids from prunus tomentosa.

BACKGROUND: In recent years, hyperuricemia and acute gouty arthritis have become increasingly common, posing a serious threat to public health. Current treatments primarily involve Western medicines with associated toxic side effects. OBJECTIVE: This study aims to investigate the therapeutic effects of total flavones from Prunus tomentosa (PTTF) on a rat model of gout and explore the mechanism of PTTF's anti-gout action through the TLR4/NF-κB signaling pathway. METHODS: We measured serum uric acid (UA), creatinine (Cr), blood urea nitrogen (BUN), tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1ß), and interleukin-6 (IL-6) levels using an enzyme-linked immunosorbent assay (ELISA). Histopathological changes were observed using HE staining, and the expression levels of relevant proteins were detected through Western blotting. RESULTS: After PTTF treatment, all indicators improved significantly. PTTF reduced blood levels of UA, Cr, BUN, IL-1ß, IL-6, and TNF-α, and decreased ankle swelling. CONCLUSIONS: PTTF may have a therapeutic effect on animal models of hyperuricemia and acute gouty arthritis by reducing serum UA levels, improving ankle swelling, and inhibiting inflammation. The primary mechanism involves the regulation of the TLR4/NF-κB signaling pathway to alleviate inflammation. Further research is needed to explore deeper mechanisms.

Chinese cherry CpMYB44-CpSPDS2 module regulates spermidine content and florescence in tobacco.

The flower bud differentiation plays a crucial role in cherry yield and quality. In a preliminary study, we revealed the promotion of spermidine (Spd) in bud differentiation and quality. However, the molecular mechanism underlying Spd regulating cherry bud differentiation remains unclear. To address this research gap, we cloned CpSPDS2, a gene that encodes Spd synthase and is highly expressed in whole flowers and pistils of the Chinese cherry (cv. 'Manaohong'). Furthermore, an overexpression vector with this gene was constructed to transform tobacco plants. The findings demonstrated that transgenic lines exhibited higher Spd content, an earlier flowering time by 6 d, and more lateral buds and flowers than wild-type lines. Additionally, yeast one-hybrid assays and two-luciferase experiments confirmed that the R2R3-MYB transcription factor (CpMYB44) directly binds to and activates the CpSPDS2 promoter transcription. It is indicated that CpMYB44 promotes Spd accumulation via regulating CpSPDS2 expression, thus accelerating the flower growth. This research provides a basis for resolving the molecular mechanism of CpSPDS2 involved in cherry bud differentiation.

Ecological drivers of flower-leaf sequences: aridity and proxies for pollinator attraction select for flowering-first in the American plums.

Across temperate forests, many tree species produce flowers before their leaves emerge. This flower-leaf phenological sequence, known as hysteranthy, is generally described as an adaptation for wind pollination. However, this explanation does not address why hysteranthy is also common in biotically pollinated taxa. We quantified flower-leaf sequence variation in the American plums (Prunus, subg. Prunus sect. Prunocerasus), a clade of insect-pollinated trees, using herbaria specimens and Bayesian hierarchical modeling. We tested two common, but rarely interrogated hypotheses - that hysteranthy confers aridity tolerance and/or pollinator visibility - by modeling the associations between hysteranthy and related traits. To understand how these phenology-trait associations were sensitive to taxonomic scale and flower-leaf sequence classification, we then extended these analyses to all Prunus species in North America. Our findings across two taxonomic levels support the hypotheses that hysteranthy may help temporally partition hydraulic demand to reduce water stress and increase pollinator visibility - thereby reducing selective pressure on inflorescence size. Our results provide foundational insights into the evolution of flower-leaf sequences in the genus Prunus, with implications for understanding these patterns in biotically pollinated plants in general. Our approach suggests a path to advance these hypotheses to other clades, but teasing out drivers fully will require new experiments.

Distinguishing features of Prunus humilis, P. japonica, P. pedunculata seeds and their adulterant based on DNA barcoding, morphological characterization, and chemical profiles.

Pruni Semen, the dried ripe seed of Prunus humilis, P. japonica, or P. pedunculata as recorded in the Chinese Pharmacopoeia, has been widely used in pharmaceutical and food industries. The adulteration of the marketed product with morphologically similar plants of the same genus has led to variable product quality and clinical effectiveness. This study systematically investigated the phylogenetic relationships, morphological traits, and chemical profiles of 37 Pruni Semen samples from planting bases, markets, and fields. DNA barcoding could successfully distinguish the genuine and counterfeit Pruni Semen, and the results indicated that there was almost no authentic Pruni Semen available in the market. The samples were divided into "big seed" (P. pedunculata and P. salicina seeds) and "small seed" (P. humilis, P. japonica, P. tomentosa, and P. avium seeds) categories based on morphology results. The notable discrepancy in the chemical characteristics of "big seed" and "small seed" was that "small seeds" were rich in flavonoids and low in amygdalin, whereas "big seeds" were the opposite. Furthermore, principal component analysis and clustered heatmap analysis verified the distinguishing features of "big seed" and "small seed" based on morphological and chemical characteristics. This study suggested that a combination of DNA barcoding and morphological and chemical characteristics can aid in the identification and quality evaluation of authentic and adulterated Pruni Semen. These findings may help standardize Pruni Semen available in the market and protect the rights and interests of customers.

Genetic dissection of the tissue-specific roles of type III effectors and phytotoxins in the pathogenicity of Pseudomonas syringae pv. syringae to cherry.

When compared with other phylogroups (PGs) of the Pseudomonas syringae species complex, P. syringae pv. syringae (Pss) strains within PG2 have a reduced repertoire of type III effectors (T3Es) but produce several phytotoxins. Effectors within the cherry pathogen Pss 9644 were grouped based on their frequency in strains from Prunus as the conserved effector locus (CEL) common to most P. syringae pathogens; a core of effectors common to PG2; a set of PRUNUS effectors common to cherry pathogens; and a FLEXIBLE set of T3Es. Pss 9644 also contains gene clusters for biosynthesis of toxins syringomycin, syringopeptin and syringolin A. After confirmation of virulence gene expression, mutants with a sequential series of T3E and toxin deletions were pathogenicity tested on wood, leaves and fruits of sweet cherry (Prunus avium) and leaves of ornamental cherry (Prunus incisa). The toxins had a key role in disease development in fruits but were less important in leaves and wood. An effectorless mutant retained some pathogenicity to fruit but not wood or leaves. Striking redundancy was observed amongst effector groups. The CEL effectors have important roles during the early stages of leaf infection and possibly acted synergistically with toxins in all tissues. Deletion of separate groups of T3Es had more effect in P. incisa than in P. avium. Mixed inocula were used to complement the toxin mutations in trans and indicated that strain mixtures may be important in the field. Our results highlight the niche-specific role of toxins in P. avium tissues and the complexity of effector redundancy in the pathogen Pss 9644.

Comparative genomics of N-acetyl-5-methoxytryptamine members in four Prunus species with insights into bud dormancy and abiotic stress responses in Prunus avium.

KEY MESSAGE: This study provides novel insights into the evolution, diversification, and functions of melatonin biosynthesis genes in Prunus species, highlighting their potential role in regulating bud dormancy and abiotic stresses. The biosynthesis of melatonin (MEL) in plants is primarily governed by enzymatic reactions involving key enzymes such as serotonin N-acetyltransferase (SNAT), tryptamine 5-hydroxylase (T5H), N-acetylserotonin methyltransferase (ASMT) and tryptophan decarboxylase (TDC). In this study, we analyzed Melatonin genes in four Prunus species such as Prunus avium (Pavi), Prunus pusilliflora (Ppus), Prunus serulata (Pser), and Prunus persica (Pper) based on comparative genomics approach. Among the four Prunus species, a total of 29 TDCs, 998 T5Hs, 16 SNATs, and 115 ASMTs within the genome of four Prunus genomes. A thorough investigation of melatonin-related genes was carried out using systematic biological methods and comparative genomics. Through phylogenetic analysis, orthologous clusters, Go enrichment, syntenic relationship, and gene duplication analysis, we discovered both similarities and variations in Melatonin genes among these Prunus species. Additionally, our study revealed the existence of unique subgroup members in the Melatonin genes of these species, which were distinct from those found in Arabidopsis genes. Furthermore, the transcriptomic expression analysis revealed the potential significance of melatonin genes in bud dormancy regulation and abiotic stresses. Our extensive results offer valuable perspectives on the evolutionary patterns, intricate expansion, and functions of PavMEL genes. Given their promising attributes, PavTDCs, PavT5H, PavNAT, and three PavASMT genes warrant in-depth exploration as prime candidates for manipulating dormancy in sweet cherry. This was done to lay the foundation for future explorations into the structural and functional aspects of these factors in Prunus species. This study offers significant insights into the functions of ASMT, SNAT, T5H, and TDC genes and sheds light on their roles in Prunus avium. Moreover, it established a robust foundation for further exploration functional characterization of melatonin genes in fruit species.

Mutations overlying the miR172 target site of TOE-type genes are prime candidate variants for the double-flower trait in mei.

Mutations affecting flower shape in many plants have been favored by human selection, and various fruit trees are also grown for ornamental purposes. Mei (Prunus mume) is a dual purpose tree originated in China well known in the Western world for its generous early blooms, often bearing double flowers. Building on the knowledge of its genomic location, a candidate gene approach was used to identify a 49 bp deletion encompassing the miR172 target site of the euAP2 gene pmTOE (PmuVar_Ch1_3490) as a prime variant linked to flower doubleness. Searching within a large dataset of genome sequencing data from Eastern germplasm collections demonstrated a tight variant-trait association, further confirmed in a panel of commercial and non-commercial varieties available in Italy. Moreover, two SNP mutations in the miR172 target site of pmPET (PmuVar_Ch1_1333) were identified in some double flower accessions. The mei orthologue of PETALOSA genes already found responsible for the phenotype in other plants suggests that independent variants may have been selected throughout mei domestication history.

Effects of almond (Armeniaca Sibirica L. Lam) polysaccharides on gut microbiota and anti-inflammatory effects on LPS-induced RAW264.7 cells.

The aim of this study was to investigate the prebiotic properties of the almond polysaccharide AP-1 on intestinal microorganisms by using an in vitro fecal fermentation method and its anti-inflammatory effect on lipopolysaccharide (LPS)-induced RAW264.7 cells. The results showed that during the in vitro fermentation of AP-1, the pH value of the fermentation broth decreased obviously, while the concentration of short-chain fatty acids (SCFAs) increased significantly, especially acetic acid and butyric acid. In genus level, the number of Clostridium and Megamonas increased markedly in the AP-1 group after 24 h of fermentation. After 48 h of fermentation, there was a noticeable increase in the number of beneficial genera Lactobacillaceae and Bifidobacteriaceae, and a considerable decrease in the number of pro-inflammatory genera. In addition, we found that AP-1 had no toxic effect on RAW264.7 cells. In the LPS-induced inflammation model of RAW264.7 cells, AP-1 could effectively inhibit the release of NO, regulate the level of reactive oxides (ROS), and effectively down-regulate the mRNA expression of TNF-α, IL-1ß, IL-6 and iNOS. In conclusion, the almond polysaccharide AP-1 may be a functional active substance aimed at promoting intestinal health and exerting anti-inflammatory effects.

Molecular characterization and evolutionary relationships of DOFs in four cherry species and functional analysis in sweet cherry.

The DOF (DNA binding with one finger) has multiple functions in plants. However, it has received little attention in the research field of cherries. In this study, the evolutionary relationship and molecular characterization of DOF in four cherry species were analyzed, revealing its expression pattern in sweet cherry. There are 23 members in Prunus avium cv. 'Tieton', 88 in Prunus cerasus, 53 in Cerasus × yedoensis, and 27 in Cerasus serrulata. Most of these genes are intron-less or non-intron, with a conserved C2-C2 domain. Due to heterozygosity and chromosomal ploidy, whole-genome duplication (WGD) events occur to varying degrees, and DOF genes are contracted during evolution. Furthermore, these genes are affected by purifying selection pressure. Under low-temperature treatment, the expression of PavDOF2 and PavDOF18 were significantly up-regulated, while that of PavDOF16 is significantly down-regulated. The expression of PavDOF9, PavDOF12, PavDOF14, PavDOF16, PavDOF17, PavDOF18, and PavDOF19 exhibits an increasing trend during flower development and varies during sweet cherry fruit development. PavDOF1, PavDOF8, PavDOF9, and PavDOF15 are localized in the nucleus but is not transcriptionally active. The findings systemically demonstrate the molecular characteristics of DOF in different cherry varieties, providing a basis for further research on the functions of these genes.

AGAMOUS-LIKE24 controls pistil number in Japanese apricot by targeting the KNOTTED1-LIKE gene KNAT2/6-a.

The formation of multi-pistil flowers reduces the yield and quality in Japanese apricot (Prunus mume). However, the molecular mechanism underlying the formation of multi-pistil flowers remains unknown. In the current study, overexpression of PmKNAT2/6-a, a class I KNOTTED1-like homeobox (KNOX) member, in Arabidopsis (Arabidopsis thaliana) resulted in a multi-pistil phenotype. Analysis of the upstream regulators of PmKNAT2/6-a showed that AGAMOUS-like 24 (PmAGL24) could directly bind to the PmKNAT2/6-a promoter and regulate its expression. PmAGL24 also interacted with Like Heterochromatin Protein 1 (PmLHP1) to recruit lysine trimethylation at position 27 on histone H3 (H3K27me3) to regulate PmKNAT2/6-a expression, which is indirectly involved in multiple pistils formation in Japanese apricot flowers. Our study reveals that the PmAGL24 transcription factor, an upstream regulator of PmKNAT2/6-a, regulates PmKNAT2/6-a expression via direct and indirect pathways and is involved in the formation of multiple pistils in Japanese apricot.

Almond Can Be Infected by Plum Pox Virus-D Isolate Penn4 and Is a Transmission-Competent Host.

Although it is currently eradicated from the United States, Plum pox virus (PPV) poses an ongoing threat to U.S. stone fruit production. Although almond (Prunus dulcis) is known to be largely resistant to PPV, there is conflicting evidence about its potential to serve as an asymptomatic reservoir host for the virus and thus serve as a potential route of entry. Here, we demonstrate that both Tuono and Texas Mission cultivars can be infected by the U.S. isolate PPV Dideron (D) Penn4 and that Tuono is a transmission-competent host, capable of serving as a source of inoculum for aphid transmission of the virus. These findings have important implications for efforts to keep PPV out of the United States and highlight the need for additional research to test the susceptibility of almond to other PPV-D isolates.

Allelic variation of PmCBF03 contributes to the altitude and temperature adaptability in Japanese apricot (Prunus mume Sieb. et Zucc.).

Japanese apricot is an important subtropical deciduous fruit tree in China, widely distributed in different altitude areas. How does it adapt to the different temperature environments in these areas? In this study, we identified a low-temperature transcription factor PmCBF03 on chromosome 7 through adaptive analysis of populations at different altitudes, which has an early termination single nucleotide polymorphism mutation. There were two different types of variation, PmCBF03A type in high-altitude areas and PmCBF03T type in low-altitude areas. PmCBF03A gene increased the survival rate, Fv/Fm values, antioxidant enzyme activity, and expression levels of antioxidant enzyme genes, and reducing electrolyte leakage and accumulation of reactive oxygen species in transgenic Arabidopsis under low temperature and freezing stress. Simultaneously, PmCBF03A gene promoted the dormancy of transgenic Arabidopsis seeds than wild-type. Biochemical analysis demonstrated that PmCBF03A directly bound to the DRE/CRT element in the promoters of the PmCOR413, PmDAM6 and PmABI5 genes, promoting their transcription and enhanced the cold resistance and dormancy of the overexpressing PmCBF03A lines. While PmCBF03T gene is unable to bind to the promoters of PmDAM6 and PmABI5 genes, leading to early release of dormancy to adapt to the problem of insufficient chilling requirement in low-altitude areas.

Plant height variation and genetic diversity between Prunus ledebouriana (Schlecht.) YY Yao and Prunus tenella Batsch based on using SSR markers in East Kazakhstan.

Background: Genetic differences between isolated endemic populations of plant species and those with widely known twin species are relevant for conserving the biological diversity of our planet's flora. Prunus ledebouriana (Schlecht.) YY Yao is an endangered and endemic species of shrub almond from central Asia. Few studies have explored this species, which is closely related and morphologically similar to the well-known Prunus tenella Batsch. In this article, we present a comparative analysis of studies of three P. ledebouriana populations and one close population of P. tenella in Eastern Kazakhstan in order to determine the particular geographic mutual replacement of the two species. Methods: The populations were collected from different ecological niches, including one steppe population near Ust-Kamenogorsk (P. tenella) and three populations (P. ledebouriana) in the mountainous area. Estimation of plant height using a t-test suggested a statistically significant difference between the populations and the two species (P < 0.0001). DNA simple sequence repeat (SSR) markers were applied to study the two species' genetic diversity and population structure. Results: A total of 19 polymorphic SSR loci were analyzed, and the results showed that the population collected in mountainous areas had a lower variation level than steppe populations. The highest level of Nei's genetic diversity index was demonstrated in the 4-UK population (0.622) of P. tenella. The lowest was recorded in population 3-KA (0.461) of P. ledebouriana, collected at the highest altitude of the four populations (2,086 meters above sea level). The total genetic variation of P. ledebouriana was distributed 73% within populations and 27% between populations. STRUCTURE results showed that two morphologically similar species diverged starting at step K = 3, with limited population mixing. The results confirmed the morphological and genetic differences between P. tenella and P. ledebouriana and described the level of genetic variation for P. ledebouriana. The study's results proved that the steppe zone and mountain altitude factor between P. tenella and isolated mountain samples of P. ledebouriana.

[Exploration of cross-cultivar group characteristics of a new cultivar of Prunus mume 'Zhizhang Guhong Chongcui'].

'Zhizhang Guhong Chongcui' is a new cultivar of Prunus mume with cross-cultivar group characteristics. It has typical characteristics of cinnabar purple cultivar group and green calyx cultivar group. It has green calyx, white flower, and light purple xylem, but the mechanism remains unclear. In order to clarify the causes of its cross-cultivar group traits, the color phenotype, anthocyanin content and the expression levels of genes related to anthocyanin synthesis pathway of 'Zhizhang Guhong Chongcui', 'Yuxi Zhusha' and 'Yuxi Bian Lü'e' were determined. It was found that the red degree of petals, sepals and fresh xylem in branches was positively correlated with the total anthocyanin content. MYBɑ1, MYB1, and bHLH3 were the key transcription factor genes that affected the redness of the three cultivars of flowers and xylem. The transcription factors further promoted the high expression of structural genes F3'H, DFR, ANS and UFGT, thereby promoting the production of red traits. Combined with phenotype, anthocyanin content and qRT-PCR results, it was speculated that the white color of petals of 'Zhizhang Guhong Chongcui' were derived from the high expression of FLS, F3'5'H, LAR and ANR genes in other branches of cyanidin synthesis pathway, and the low expression of GST gene. The green color of sepals might be originated from the relatively low expression of F3'H, DFR and ANS genes. The red color of xylem might be derived from the high expression of ANS and UFGT genes. This study made a preliminary explanation for the characteristics of the cross-cultivar group of 'Zhizhang Guhong Chongcui', and provided a reference for molecular breeding of flower color and xylem color of Prunus mume.