Synthetic polyploidization induces enhanced phytochemical profile and biological activities in Thymus vulgaris L. essential oil.

Essential oil from Thymus vulgaris L. has valuable therapeutic potential that is highly desired in pharmaceutical, food, and cosmetic industries. Considering these advantages and the rising market demand, induced polyploids were obtained using oryzalin to enhance essential oil yield. However, their therapeutic values were unexplored. So, this study aims to assess the phytochemical content, and antimicrobial, antioxidant, and anti-inflammatory activities of tetraploid and diploid thyme essential oils. Induced tetraploids had 41.11% higher essential oil yield with enhanced thymol and γ-terpinene content than diploid. Tetraploids exhibited higher antibacterial activity against all tested microorganisms. Similarly, in DPPH radical scavenging assay tetraploid essential oil was more potent with half-maximal inhibitory doses (IC50) of 180.03 µg/mL (40.05 µg TE/mg) than diploid with IC50 > 512 µg/mL (12.68 µg TE/mg). Tetraploids exhibited more effective inhibition of in vitro catalytic activity of pro-inflammatory enzyme cyclooxygenase-2 (COX-2) than diploids at 50 µg/mL concentration. Furthermore, molecular docking revealed higher binding affinity of thymol and γ-terpinene towards tested protein receptors, which explained enhanced bioactivity of tetraploid essential oil. In conclusion, these results suggest that synthetic polyploidization using oryzalin could effectively enhance the quality and quantity of secondary metabolites and can develop more efficient essential oil-based commercial products using this induced genotype.

Genome-Wide Identification of LOX Gene Family and Its Expression Analysis under Abiotic Stress in Potato (Solanum tuberosum L.).

The lipoxygenases (LOXs) are non-heme iron-containing dioxygenases that play an important role in plant growth and defense responses. There is scarce knowledge regarding the LOX gene family members and their involvement in biotic and abiotic stresses in potato. In this study, a total of 17 gene family members (StLOXs) in potato were identified and clustered into three subfamilies: 9-LOX type I, 13-LOX type I, and 13-LOX type II, with eleven, one, and five members in each subfamily based on phylogenetic analysis. By exploiting the RNA-seq data in the Potato Genome Sequencing Consortium (PGSC) database, the tissue-specific expressed and stress-responsive StLOX genes in double-monoploid (DM) potato were obtained. Furthermore, six candidate StLOX genes that might participate in drought and salt response were determined via qPCR analysis in tetraploid potato cultivars under NaCl and PEG treatment. Finally, the involvement in salt stress response of two StLOX genes, which were significantly up-regulated in both DM and tetraploid potato under NaCl and PEG treatment, was confirmed via heterologous expression in yeast under salt treatment. Our comprehensive analysis of the StLOX family provides a theoretical basis for the potential biological functions of StLOXs in the adaptation mechanisms of potato to stress conditions.

Whole-genome sequencing of tetraploid potato varieties reveals different strategies for drought tolerance.

Climate changes leading to increasingly longer seasonal drought periods in large parts of the world increase the necessity for breeding drought-tolerant crops. Cultivated potato (Solanum tuberosum), the third most important vegetable crop worldwide, is regarded as drought-sensitive due to its shallow root architecture. Two German tetraploid potato cultivars differing in drought tolerance and their F1-progeny were evaluated under various drought scenarios. Bulked segregant analyses were combined with whole-genome sequencing (BSA-Seq) using contrasting bulks of drought-tolerant and drought-sensitive F1-clones. Applying QTLseqr, 15 QTLs comprising 588,983 single nucleotide polymorphisms (SNPs) in 2325 genes associated with drought stress tolerance were identified. SeqSNP analyses in an association panel of 34 mostly starch potato varieties using 1-8 SNPs for each of 188 selected genes narrowed the number of candidate genes down to 10. In addition, ent-kaurene synthase B was the only gene present under QTL 10. Eight of the identified genes (StABP1, StBRI1, StKS, StLEA, StPKSP1, StPKSP2, StYAB5, and StZOG1) address plant development, the other three genes (StFATA, StHGD and StSYP) contribute to plant protection under drought stress. Allelic variation in these genes might be explored in future breeding for drought-tolerant potato varieties.

Persistence of Chronic Lymphocytic Leukemia Stem-like Populations under Simultaneous In Vitro Treatment with Curcumin, Fludarabine, and Ibrutinib: Implications for Therapy Resistance.

Leukemic stem cells (LSCs) possess similar characteristics to normal hematopoietic stem cells, including self-renewal capacity, quiescence, ability to initiate leukemia, and drug resistance. These cells play a significant role in leukemia relapse, persisting even after apparent remission. LSCs were first described in 1994 by Lapidot et al. Although they have been extensively studied in acute leukemia, more LSC research is still needed in chronic lymphocytic leukemia (CLL) to understand if reduced apoptosis in mature cells should still be considered as the major cause of this disease. Here, we provide new evidence suggesting the existence of stem-like cell populations in CLL, which may help to understand the disease as well as to develop effective treatments. In this study, we identified a potential leukemic stem cell subpopulation using the tetraploid CLL cell line I83. This subpopulation is characterized by diploid cells that were capable of generating the I83 tetraploid population. Furthermore, we adapted a novel flow cytometry analysis protocol to detect CLL subpopulations with stem cell properties in peripheral blood samples and primary cultures from CLL patients. These cells were identified by their co-expression of CD19 and CD5, characteristic markers of CLL cells. As previously described, increased alkaline phosphatase (ALP) activity is indicative of stemness and pluripotency. Moreover, we used this method to investigate the potential synergistic effect of curcumin in combination with fludarabine and ibrutinib to deplete this subpopulation. Our results confirmed the effectiveness of this ALP-based analysis protocol in detecting and monitoring leukemic stem-like cells in CLL. This analysis also identified limitations in eradicating these populations using in vitro testing. Furthermore, our findings demonstrated that curcumin significantly enhanced the effects of fludarabine and ibrutinib on the leukemic fraction, exhibiting synergistic effects (combination drug index, CDI 0.97 and 0.37, respectively). Our results lend support to the existence of potential stem-like populations in CLL cell lines, and to the idea that curcumin could serve as an effective adjuvant in therapies aimed at eliminating these populations and improving treatment efficacy.

Temporal profiling of physiological, histological, and transcriptomic dissection during auxin-induced adventitious root formation in tetraploid Robinia pseudoacacia micro-cuttings.

MAIN CONCLUSION: Optimal levels of indole-3-butyric acid (IBA) applied at the stem base promote adventitious root (AR) initiation and primordia formation, thus promoting the rooting of leafy micro-cuttings of tetraploid Robinia pseudoacacia. Tetraploid Robinia pseudoacacia L. is a widely cultivated tree in most regions of China that has a hard-rooting capability, propagated by stem cuttings. This study utilizes histological, physiological, and transcriptomic approaches to explore how root primordia are induced after indole butyric acid (IBA) treatment of micro-cuttings. IBA application promoted cell divisions in some cells within the vasculature, showing subcellular features associated with adventitious root (AR) founder cells. The anatomical structure explicitly showed that AR initiated from the cambium layer and instigate the inducible development of AR primordia. Meanwhile, the hormone data showed that similar to that of indole-3-acetic acid, the contents of trans-zeatin and abscisic acid peaked at early stages of AR formation and increased gradually in primordia formation across the subsequent stages, suggesting their indispensable roles in AR induction. On the contrary, 24-epibrassinolide roughly maintained at extremely high levels during primordium initiation thoroughly, indicating its presence was involved in cell-specific reorganization during AR development. Furthermore, antioxidant activities transiently increased in the basal region of micro-cuttings and may serve as biochemical indicators for distinct rooting phases, potentially aiding in AR formation. Transcriptomic analysis during the early stages of root formation shows significant downregulation of the abscisic acid and jasmonate signaling pathways, while ethylene and cytokinin signaling seems upregulated. Network analysis of genes involved in carbon metabolism and photosynthesis indicates that the basal region of the micro-cuttings undergoes rapid reprogramming, which results in the breakdown of sugars into pyruvate. This pyruvate is then utilized to fuel the tricarboxylic acid cycle, thereby sustaining growth through aerobic respiration. Collectively, our findings provide a time-course morphophysiological dissection and also suggest the regulatory role of a conserved auxin module in AR development in these species.

Transcriptomic and Phenotypic Analyses Reveal the Molecular Mechanism of Dwarfing in Tetraploid Robinia pseudoacacia L.

Polyploid breeding techniques aid in the cultivation of new forestry cultivars, thus expanding the suite of strategies for the improvement of arboreal traits and innovation within the field of forestry. Compared to diploid Robinia pseudoacacia L. (black locust) 'D26-5①' (2×), its dwarfed homologous tetraploid 'D26-5②' (4×) variety has better application prospects in garden vegetation guardrails and urban landscape. However, the molecular mechanism of the generation and growth of this dwarf variety is still unclear. Here, plant growth and development as well as histological differences between the diploid and its autotetraploid were investigated. Levels of endogenous hormones at three different developmental stages (20, 40, and 70 days) of 2× and homologous 4× tissue culture plantlets were assessed, and it was found that the brassinosteroid (BR) contents of the former were significantly higher than the latter. Transcriptome sequencing data analysis of 2× and homologous 4× showed that differentially expressed genes (DEGs) were significantly enriched in plant hormone synthesis and signal transduction, sugar and starch metabolism, and the plant circadian rhythm pathway, which are closely related to plant growth and development. Therefore, these biological pathways may be important regulatory pathways leading to dwarfism and slow growth in tetraploids. Additionally, utilizing weighted gene coexpression network analysis (WGCNA), we identified three crucial differentially expressed genes (DEGs)-PRR5, CYP450, and SPA1-that potentially underlie the observed ploidy variation. This study provides a new reference for the molecular mechanism of dwarfism in dwarfed autotetraploid black locusts. Collectively, our results of metabolite analysis and comparative transcriptomics confirm that plant hormone signaling and the circadian rhythm pathway result in dwarfism in black locusts.

Haplotype-resolved assembly of a tetraploid potato genome using long reads and low-depth offspring data.

Potato is one of the world's major staple crops, and like many important crop plants, it has a polyploid genome. Polyploid haplotype assembly poses a major computational challenge. We introduce a novel strategy for the assembly of polyploid genomes and present an assembly of the autotetraploid potato cultivar Altus. Our method uses low-depth sequencing data from an offspring population to achieve chromosomal clustering and haplotype phasing on the assembly graph. Our approach generates high-quality assemblies of individual chromosomes with haplotype-specific sequence resolution of whole chromosome arms and can be applied in common breeding scenarios where collections of offspring are available.

Multienvironment genomic prediction in tetraploid potato.

Multienvironment genomic prediction was applied to tetraploid potato using 147 potato varieties, tested for 2 years, in 3 locations representative of 3 distinct regions in Europe. Different prediction scenarios were investigated to help breeders predict genotypic performance in the regions from one year to the next, for genotypes that were tested this year (scenario 1), as well as new genotypes (scenario 3). In scenario 2, we predicted new genotypes for any one of the 6 trials, using all the information that is available. The choice of prediction model required assessment of the variance-covariance matrix in a mixed model that takes into account heterogeneity of genetic variances and correlations. This was done for each analyzed trait (tuber weight, tuber length, and dry matter) where examples of both limited and higher degrees of heterogeneity was observed. This explains why dry matter did not need complex multienvironment modeling to combine environments and increase prediction ability, while prediction in tuber weight, improved only when models were flexible enough to capture the heterogeneous variances and covariances between environments. We also found that the prediction abilities in a target trial condition decreased, if trials with a low genetic correlation to the target were included when training the model. Genomic prediction in tetraploid potato can work once there is clarity about the prediction scenario, a suitable training set is created, and a multienvironment prediction model is chosen based on the patterns of G×E indicated by the genetic variances and covariances.

Induction of tetraploids in Paper Mulberry (Broussonetia papyrifera (L.) L'Hér. ex Vent.) by colchicine.

BACKGROUND: Broussonetia papyrifera (L.) L'Hér. ex Vent. has the characteristics of strong stress resistance, high crude protein content, and pruning tolerance. It is an ecological, economic, and medicinal plant. Polyploid plants usually perform better than their corresponding diploid plants in terms of nutrients, active substances, and stress resistance. RESULTS: In this study, the leaves, calli, and seeds of diploid B. papyrifera were used for tetraploid induction by colchicine. The induction effect of colchicine on B. papyrifera was summarized through the early morphology, chromosome count and flow cytometry. It was concluded that the best induction effect (18.6%) was obtained when the leaves of B. papyrifera were treated in liquid MS (Murashige and Skoog) medium containing 450 mg·L-1 colchicine for 3 d. The comparative analysis of the growth characteristics of diploid and tetraploid B. papyrifera showed that tetraploid B. papyrifera has larger ground diameter, larger stomata, thicker palisade tissue and thicker sponge tissue than diploid B. papyrifera. In addition, the measurement of photosynthetic features also showed that tetraploids had higher chlorophyll content and higher photosynthetic rates. CONCLUSION: This study showed that tetraploid B. papyrifera could be obtained by treating leaves, callus and seeds with liquid and solid colchicine, but the induction efficiency was different. Moreover, there were differences in stomata, leaf cell structure and photosynthetic features between tetraploid B. papyrifera and its corresponding diploid. The induced tetraploid B. papyrifera can provide a technical basis and breeding material for the creation of B. papyrifera germplasm resources in the future.

Cytological observation and RNA-seq analysis reveal novel miRNAs high expression associated with the pollen fertility of neo-tetraploid rice.

BACKGROUND: Neo-tetraploid rice lines exhibit high fertility and strong heterosis and harbor novel specific alleles, which are useful germplasm for polyploid rice breeding. However, the mechanism of the fertility associated with miRNAs remains unknown. In this study, a neo-tetraploid rice line, termed Huaduo21 (H21), was used. Cytological observation and RNA-sequencing were employed to identify the fertility-related miRNAs in neo-tetraploid rice. RESULTS: H21 showed high pollen fertility (88.08%), a lower percentage of the pollen mother cell (PMC) abnormalities, and lower abnormalities during double fertilization and embryogenesis compared with autotetraploid rice. A total of 166 non-additive miRNAs and 3108 non-additive genes were detected between H21 and its parents. GO and KEGG analysis of non-additive genes revealed significant enrichments in the DNA replication, Chromosome and associated proteins, and Replication and repair pathways. Comprehensive multi-omics analysis identified 32 pairs of miRNA/target that were associated with the fertility in H21. Of these, osa-miR408-3p and osa-miR528-5p displayed high expression patterns, targeted the phytocyanin genes, and were associated with high pollen fertility. Suppression of osa-miR528-5p in Huaduo1 resulted in a low seed set and a decrease in the number of grains. Moreover, transgenic analysis implied that osa-MIR397b-p3, osa-miR5492, and osa-MIR5495-p5 might participate in the fertility of H21. CONCLUSION: Taken together, the regulation network of fertility-related miRNAs-targets pairs might contribute to the high seed setting in neo-tetraploid rice. These findings enhance our understanding of the regulatory mechanisms of pollen fertility associated with miRNAs in neo-tetraploid rice.

Genetic architecture of tuber-bound free amino acids in potato and effect of growing environment on the amino acid content.

Free amino acids in potato tubers contribute to their nutritional value and processing quality. Exploring the natural variation in their accumulation in tubers across diverse genetic backgrounds is critical to potato breeding programs aiming to enhance or partition their distribution effectively. This study assessed variation in the tuber-bound free amino acids in a diversity panel of tetraploid potato clones developed and maintained by the Texas A&M Potato Breeding Program to explore their genetic basis and to obtain genomic-estimated breeding values for applied breeding purposes. Free amino acids content was evaluated in tubers of 217 tetraploid potato clones collected from Dalhart, Texas in 2019 and 2020, and Springlake, Texas in 2020. Most tuber amino acids were not affected by growing location, except histidine and proline, which were significantly lower (- 59.0%) and higher (+ 129.0%), respectively, at Springlake, Texas (a location that regularly suffers from abiotic stresses, mainly high-temperature stress). Single nucleotide polymorphism markers were used for genome-wide association studies and genomic selection of clones based on amino acid content. Most amino acids showed significant variations among potato clones and moderate to high heritabilities. Principal component analysis separated fresh from processing potato market classes based on amino acids distribution patterns. Genome-wide association studies discovered 33 QTL associated with 13 free amino acids. Genomic-estimated breeding values were calculated and are recommended for practical potato breeding applications to select parents and advance clones with the desired free amino acid content.

Genome assembly of two diploid and one auto-tetraploid Cyclocarya paliurus genomes.

Cyclocarya paliurus, an endemic species in the genus Juglandaceae with the character of heterodichogamy, is one of triterpene-rich medicinal plants in China. To uncover the genetic mechanisms behind the special characteristics, we sequenced the genomes of two diploid (protandry, PA-dip and protogyny, PG-dip) and one auto-tetraploid (PA-tetra) C. paliurus genomes. Based on 134.9 (~225x), 75.5 (~125x) and 271.8 Gb (~226x) subreads of PacBio platform sequencing data, we assembled 586.62 Mb (contig N50 = 1.9 Mb), 583.45 Mb (contig N50 = 1.4 Mb), and 2.38 Gb (contig N50 = 430.9 kb) for PA-dip, PG-dip and PA-tetra genome, respectively. Furthermore, 543.53, 553.87, and 2168.65 Mb in PA-dip, PG-dip, and PA-tetra, were respectively anchored to 16, 16, and 64 pseudo-chromosomes using over 65.4 Gb (~109x), 68 Gb (~113x), and 264 (~220x) Hi-C sequencing data. Annotation of PA-dip, PG-dip, and PA-tetra genome assembly identified 34,699, 35,221, and 34,633 protein-coding genes (90,752 gene models) or allele-defined genes, respectively. In addition, 45 accessions from nine locations were re-sequenced, and more than 10 × coverage reads were generated.

Genome-material costs and functional trade-offs in the autopolyploid Solidago gigantea (giant goldenrod) series.

PREMISE: Increased genome-material costs of N and P atoms inherent to organisms with larger genomes have been proposed to limit growth under nutrient scarcities and to promote growth under nutrient enrichments. Such responsiveness may reflect a nutrient-dependent diploid versus polyploid advantage that could have vast ecological and evolutionary implications, but direct evidence that material costs increase with ploidy level and/or influence cytotype-dependent growth, metabolic, and/or resource-use trade-offs is limited. METHODS: We grew diploid, autotetraploid, and autohexaploid Solidago gigantea plants with one of four ambient or enriched N:P ratios and measured traits related to material costs, primary and secondary metabolism, and resource-use. RESULTS: Relative to diploids, polyploids invested more N and P into cells, and tetraploids grew more with N enrichments, suggesting that material costs increase with ploidy level. Polyploids also generally exhibited strategies that could minimize material-cost constraints over both long (reduced monoploid genome size) and short (more extreme transcriptome downsizing, reduced photosynthesis rates and terpene concentrations, enhanced N-use efficiencies) evolutionary time periods. Furthermore, polyploids had lower transpiration rates but higher water-use efficiencies than diploids, both of which were more pronounced under nutrient-limiting conditions. CONCLUSIONS: N and P material costs increase with ploidy level, but material-cost constraints might be lessened by resource allocation/investment mechanisms that can also alter ecological dynamics and selection. Our results enhance mechanistic understanding of how global increases in nutrients might provide a release from material-cost constraints in polyploids that could impact ploidy (or genome-size)-specific performances, cytogeographic patterning, and multispecies community structuring.

Canonical and Alternative Auxin Signaling Systems in Mono-, Di-, and Tetraploid Potatoes.

It has long been known that the phytohormone auxin plays a promoting role in tuber formation and stress tolerance in potatoes. Our study aimed to identify and characterize the complete sets of auxin-related genes that presumably constitute the entire auxin signaling system in potato (Solanum tuberosum L.). The corresponding genes were retrieved from sequenced genomes of the doubled monoploid S. tuberosum DM1-3-516-R44 (DM) of the Phureja group, the heterozygous diploid line RH89-039-16 (RH), and the autotetraploid cultivar Otava. Both canonical and noncanonical auxin signaling pathways were considered. Phylogenetic and domain analyses of deduced proteins were supplemented by expression profiling and 3D molecular modeling. The canonical and ABP1-mediated pathways of auxin signaling appeared to be well conserved. The total number of potato genes/proteins presumably involved in canonical auxin signaling is 46 and 108 in monoploid DM and tetraploid Otava, respectively. Among the studied potatoes, spectra of expressed genes obviously associated with auxin signaling were partly cultivar-specific and quite different from analogous spectrum in Arabidopsis. Most of the noncanonical pathways found in Arabidopsis appeared to have low probability in potato. This was equally true for all cultivars used irrespective of their ploidy. Thus, some important features of the (noncanonical) auxin signaling pathways may be variable and species-specific.

Exposure to Pfaffia glomerata causes oxidative stress and triggers hepatic changes.

Medicinal plant species are genetically engineered to obtain higher production of biomass and specific secondary metabolites, which can be used in the pharmaceutical industry. The aim of the present study was to evaluate the effect of Pfaffia glomerata (Spreng.) Pedersen tetraploid hydroalcoholic extract on the liver of adult Swiss mice. The extract was prepared from the plant roots and given to the animals by gavage, for 42 days. The experimental groups were treated with water (control), Pfaffia glomerata tetraploid hydroalcoholic extract (100, 200 and 400 mg/kg) and Pfaffia glomerata tetraploid hydroalcoholic extract discontinuously (200 mg/kg). The last group received the extract every 3 days, for 42 days. The oxidative status, mineral dynamics, and cell viability were analysed. The liver weight and the number of viable hepatocytes were reduced, despite the increased cell's number. Increased levels of malondialdehyde and nitric oxide, and changes in iron, copper, zinc, potassium, manganese and sodium levels were observed. aspartate aminotransferase levels were increased while alanine aminotransferase levels were decreased due to BGEt intake. Our results showed that BGEt induced alterations of oxidative stress biomarkers leading to liver injury, which was associated with a reduction in the number of hepatocytes.

Comparative Transcriptome Analysis Revealed Key Genes Regulating Gossypol Synthesis in Tetraploid Cultivated Cotton.

Tetraploid cultivated cotton (Gossypium spp.) produces cottonseeds rich in protein and oil. Gossypol and related terpenoids, stored in the pigment glands of cottonseeds, are toxic to human beings and monogastric animals. However, a comprehensive understanding of the genetic basis of gossypol and gland formation is still lacking. We performed a comprehensive transcriptome analysis of four glanded versus two glandless tetraploid cultivars distributed in Gossypium hirsutum and Gossypium barbadense. A weighted gene co-expression network analysis (WGCNA) based on 431 common differentially expressed genes (DEGs) uncovered a candidate module that was strongly associated with the reduction in or disappearance of gossypol and pigment glands. Further, the co-expression network helped us to focus on 29 hub genes, which played key roles in the regulation of related genes in the candidate module. The present study contributes to our understanding of the genetic basis of gossypol and gland formation and serves as a rich potential source for breeding cotton cultivars with gossypol-rich plants and gossypol-free cottonseed, which is beneficial for improving food safety, environmental protection, and economic gains of tetraploid cultivated cotton.

Inbreeding Effects on the Performance and Genomic Prediction for Polysomic Tetraploid Potato Offspring Grown at High Nordic Latitudes.

Inbreeding depression (ID) is caused by increased homozygosity in the offspring after selfing. Although the self-compatible, highly heterozygous, tetrasomic polyploid potato (Solanum tuberosum L.) suffers from ID, some argue that the potential genetic gains from using inbred lines in a sexual propagation system of potato are too large to be ignored. The aim of this research was to assess the effects of inbreeding on potato offspring performance under a high latitude and the accuracy of the genomic prediction of breeding values (GEBVs) for further use in selection. Four inbred (S1) and two hybrid (F1) offspring and their parents (S0) were used in the experiment, with a field layout of an augmented design with the four S0 replicated in nine incomplete blocks comprising 100, four-plant plots at Umeå (63°49'30″ N 20°15'50″ E), Sweden. S0 was significantly (p < 0.01) better than both S1 and F1 offspring for tuber weight (total and according to five grading sizes), tuber shape and size uniformity, tuber eye depth and reducing sugars in the tuber flesh, while F1 was significantly (p < 0.01) better than S1 for all tuber weight and uniformity traits. Some F1 hybrid offspring (15-19%) had better total tuber yield than the best-performing parent. The GEBV accuracy ranged from -0.3928 to 0.4436. Overall, tuber shape uniformity had the highest GEBV accuracy, while tuber weight traits exhibited the lowest accuracy. The F1 full sib's GEBV accuracy was higher, on average, than that of S1. Genomic prediction may facilitate eliminating undesired inbred or hybrid offspring for further use in the genetic betterment of potato.

Terpenoid VOC profiles and functional characterization of terpene synthases in diploid and tetraploid cytotypes of Chrysanthemum indicum L.

Chrysanthemum indicum L. is a valuable medicinal plant with diploid and tetraploid forms that are widely distributed in central and southern China, and it contains abundant volatile organic compounds (VOCs). Despite the discovery of some terpene synthase (TPS) in C. indicum (i.e., CiTPS) in previous studies, many TPSs and their corresponding terpene biosynthesis pathways have yet to be discovered. In the present study, terpenoid VOCs in different tissues from two cytotypes of C. indicum were analyzed. We identified 52 types of terpenoid VOCs and systematically investigated the content and distribution of these compounds in various tissues. The two cytotypes of C. indicum exhibited different volatile terpenoid profiles. The content of monoterpenes and sesquiterpenes in the two cytotypes showed an opposite trend. In addition, four full-length candidate TPSs (named CiTPS5-8) were cloned from Ci-GD4x, and their homologous TPS genes were screened based on the genome data of Ci-HB2x. These eight TPSs displayed various tissue expression patterns and were discovered to produce 22 terpenoids, 5 of which are monoterpenes and 17 are sesquiterpenes. We further proposed corresponding terpene synthesis pathways, which can enable the establishment of an understanding of the volatile terpenoid profiles of C. indicum with different cytotypes. This knowledge may provide a further understanding of germplasm in C. indicum and may be useful for biotechnology applications of Chrysanthemum plants.

Pfaffia glomerata polyploid accession compromises male fertility and fetal development.

ETHNOPHARMACOLOGICAL RELEVANCE: Pfaffia glomerata (Spreng.) Pedersen has traditionally been used as a tonic and a stimulant by the Brazilian population. It shows higher biomass accumulation and production of secondary compounds, such as the phytosterol 20-hydroxyecdysone. AIMS: The present study aimed to evaluate the effects of the hydroalcoholic extract of the root of tetraploid P. glomerata (BGEt) on testicular parenchyma, and its implications on fertility. MATERIAL AND METHODS: Adult Swiss mice were divided as: control (water) and sildenafil citrate (7 mg/kg), BGEt at 100, 200, and 400 mg/kg, and BGEtD 200 mg/kg (treated with BGE every three days). Males (n = 4/group) were mated with normal untreated adult females to assess fertility rates, while other animals (n = 6/group) were euthanized for testis, epididymis, and oxidative stress analyses. RESULTS: Increase in tubule diameter and epithelium height in the discontinuous group, in addition to an increase in the proportion of tubules with moderate pathologies was observed. The pre-implantation loss was lower in all treated groups. The post-implantation loss was significantly increased in all treated groups, except for the lowest BGEt dose. BGEt intake caused a decrease in daily sperm production, along with the number and quality of sperm in the epididymis. Changes were observed in protein carbonylation and hydrogen peroxide and nitric oxide levels, characterizing oxidative stress. CONCLUSIONS: The hydroalcoholic extract of P. glomerata tetraploid altered sperm and testicular parameters, compromising embryonic development after implantation.

Regenerative plantlets with improved agronomic characteristics caused by anther culture of tetraploid potato (Solanum tuberosum L.).

Objective: As the primary means of plant-induced haploid, anther culture is of great significance in quickly obtaining pure lines and significantly shortening the potato breeding cycle. Nevertheless, the methods of anther culture of tetraploid potato were still not well established. Methods: In this study, 16 potato cultivars (lines) were used for anther culture in vitro. The corresponding relation between the different development stages of microspores and the external morphology of buds was investigated. A highly-efficient anther culture system of tetraploid potatoes was established. Results: It was shown in the results that the combined use of 0.5 mg/L 1-Naphthylacetic acid (NAA), 1.0 mg/L 2,4-Dichlorophenoxyacetic acid (2,4-D), and 1.0 mg/L Kinetin (KT) was the ideal choice of hormone pairing for anther callus. Ten of the 16 potato cultivars examined could be induced callus with their respective anthers, and the induction rate ranged from 4.44% to 22.67% using this hormone combination. According to the outcome from the orthogonal design experiments of four kinds of appendages, we found that the medium with sucrose (40 g/L), AgNO3 (30 mg/L), activated carbon (3 g/L), potato extract (200 g/L) had a promotive induction effect on the anther callus. In contrast, adding 1 mg/L Zeatin (ZT) effectively facilitated callus differentiation. Conclusion: Finally, 201 anther culture plantlets were differentiated from 10 potato cultivars. Among these, Qingshu 168 and Ningshu 15 had higher efficiency than anther culture. After identification by flow cytometry and fluorescence in situ hybridization, 10 haploid plantlets (5%), 177 tetraploids (88%), and 14 octoploids (7%) were obtained. Some premium anther-cultured plantlets were further selected by morphological and agronomic comparison. Our findings provide important guidance for potato ploidy breeding.