Functional Validation of Different Alternative Splicing Variants of the Chrysanthemum lavandulifolium ClNUM1 Gene in Tobacco.

The Asteraceae are widely distributed throughout the world, with diverse functions and large genomes. Many of these genes remain undiscovered and unstudied. In this study, we discovered a new gene ClNUM1 in Chrysanthemum lavandulifolium and studied its function. In this study, bioinformatics, RT-qPCR, paraffin sectioning, and tobacco transgenics were utilized to bioinformatically analyze and functionally study the three variable splice variants of the unknown gene ClNUM1 cloned from C. lavandulifolium. The results showed that ClNUM1.1 and ClNUM1.2 had selective 3' splicing and selective 5' splicing, and ClNUM1.3 had selective 5' splicing. When the corresponding transgenic tobacco plants were subjected to abiotic stress treatment, in the tobacco seedlings, the ClNUM1.1 gene and the ClNUM1.2 gene enhanced salt and low-temperature tolerance and the ClNUM1.3 gene enhanced low-temperature tolerance; in mature tobacco plants, the ClNUM1.1 gene was able to enhance salt and low-temperature tolerance, and the ClNUM1.2 and ClNUM1.3 genes were able to enhance low-temperature tolerance. In summary, there are differences in the functions of the different splice variants and the different seedling stages of transgenic tobacco, but all of them enhanced the resistance of tobacco to a certain extent. The analysis and functional characterization of the ClNUM1 gene provided new potential genes and research directions for abiotic resistance breeding in Chrysanthemum.

Synthesis of graphene quantum dots and their applications in drug delivery.

This review focuses on the recent advances in the synthesis of graphene quantum dots (GQDs) and their applications in drug delivery. To give a brief understanding about the preparation of GQDs, recent advances in methods of GQDs synthesis are first presented. Afterwards, various drug delivery-release modes of GQDs-based drug delivery systems such as EPR-pH delivery-release mode, ligand-pH delivery-release mode, EPR-Photothermal delivery-Release mode, and Core/Shell-photothermal/magnetic thermal delivery-release mode are reviewed. Finally, the current challenges and the prospective application of GQDs in drug delivery are discussed.

The study of the characteristics of the secondary flowering of Cerasus subhirtella 'Autumnalis'.

The short flowering period of ornamental cherry trees is the main factor limiting their use in gardens. Determining the secondary flowering characteristics of ornamental cherry trees is required to prolong their flowering period. In this experiment, Cerasus subhirtella 'Autumnalis' was used as the experimental material. The phenological differences in their annual growth cycle were observed using the BBCH coding system. The cooling requirements of the flower buds were evaluated by the chilling hours model (temperature between 0 and 7.2 °C) and the Utah model. The expression of the core gene involved in bud dormancy regulation DAM (dormancy-associated MADS-box) from the completion of flower bud differentiation in one year until the following year was measured by performing real-time fluorescence-based quantitative PCR. The results showed that the flowering duration of C. subhirtella 'Autumnalis' from November to December was longer than that of C. yedoensis 'Somei Yoshino', which was from March to April. The progress from seed bud-break to flower bud opening took about 10 days for C. subhirtella 'Autumnalis', while the same stage for C. yedoensis 'Somei Yoshino' took around 20 days. Additionally, the flower buds of C. subhirtella 'Autumnalis' needed only the chilling temperature unit of 54.08 to satisfy the chilling requirement, while C. yedoensis 'Somei Yoshino' required a chilling temperature unit of 596.75. After the completion of flower bud differentiation, during low-temperature storage, the expression of DAM4 and DAM5 genes first increased and then decreased, whereas, the expression of the DAM6 gene continued to decrease, and the expression of DAM4, DAM5, and DAM6 in C. yedoensis 'Somei Yoshino' increased rapidly and was maintained at a high level. This showed that the upregulation of the expression of the DAM4, DAM5, and DAM6 genes can inhibit the flower bud germination of Cherry Blossom. The relative expression of the DAM gene of C. subhirtella 'Autumnalis' was significantly lower than that of the DAM gene of C. yedoensis 'Somei Yoshino' from the end of October to the beginning of December, leading to its secondary flowering in autumn. These results might elucidate why the flower buds of C. subhirtella 'Autumnalis' can break their internal dormancy and bloom in the autumn and then again in the following year. Our findings might provide a reference for conducting further studies on the mechanisms of secondary flowering and bud dormancy in cherries.

Development of Flowers Buds and Mixed Buds in the Dichasial Inflorescence of Geranium koreanum Kom. (Geraniaceae).

Flower bud differentiation is of great significance for understanding plant evolution and ecological adaptability. The development of flower buds and mixed buds in the dichasial inflorescence of Geranium koreanum was described in this paper. The morphogenesis, surface structure, and organ morphology at different growth stages of G. koreanum buds were examined in detail using scanning electron microscope and stereo microscope. The development of mixed buds started from the flattened apical meristem. The stipule and leaf primordia arose first. Subsequently, the hemispherical meristem was divided into two hemispheres, forming a terminal bud and floral bud primordia, followed by lateral bud differentiation. The formation of the terminal and lateral buds of G. koreanum was sequential and their differentiation positions were also different. The floral bud primordia would develop into two flower units and four bracts. The primordia of a flower bud first formed the sepal primordia, then the stamen and petal primordia, and finally the pistil primordia. Compared to the stamen primordia, the growth of the petal primordia was slower. Finally, all organs, especially the petals and pistil, grew rapidly. When the pistil and petals exceeded the stamens and the petals changed color, the flower bud was ready to bloom.

Alternative Splicing for Leucanthemella linearis NST1 Contributes to Variable Abiotic Stress Resistance in Transgenic Tobacco.

Leucanthemella linearis is a marsh plant in the family Compositae. It has good water and moisture resistance and ornamental properties, which makes it one of the important materials for chrysanthemum breeding and genetic improvement. The NST1 (NAC secondary wall enhancement factor 1) gene is associated with the thickening of the secondary walls of fiber cells in the plant ducts and the secondary xylem and plays an important role in plant stress resistance. In this study, two variable spliceosomes of the NST1 gene were identified from a chrysanthemum plant by using bioinformatics, qRT-PCR, transgene, and paraffin section methods to explore the molecular mechanism of the variable splicing of NST1 under abiotic stress. The results show that only three amino acids were found to be different between the two LlNST1 variants. After being treated with salt, drought, and low temperatures, analysis of the expression levels of the LlNST1 and LlNST1.1 genes in Ll showed that LlNST1.1 could respond to low temperatures and salt stress and had a weak response to drought stress. However, the expression level of LlNST1 under the three treatments was lower than that of LlNST1.1. LlNST1 transgenic tobacco showed increased saline-alkali resistance and low-temperature resistance at the seedling stage. LlNST1.1 transgenic tobacco also showed enhanced saline-alkali resistance and drought resistance at the seedling stage. In conclusion, the functions of the two variable spliceosomes of the NST1 gene are very different under abiotic stress. Therefore, this study verified the function of the variable spliceosome of NST1 and improved the stress resistance of the chrysanthemum plant under examination by regulating the expression of the NST protein, which lays a material foundation for the improvement of plant stress resistance materials and has important significance for the study of the resistance of chrysanthemum plants to abiotic stress.

Directional Osteo-Differentiation Effect of hADSCs on Nanotopographical Self-Assembled Polystyrene Nanopit Surfaces.

INTRODUCTION: Cells exhibit high sensitivity and a diverse response to the nanotopography of the extracellular matrix, thereby endowing materials with instructive performances formerly reserved for growth factors. This finding leads to opportunities for improvement. However, the interplay between the topographical surface and cell behaviors remains incompletely understood. METHODS: In the present study, we showed nanosurfaces with various dimensions of nanopits (200-750 nm) fabricated by self-assembling polystyrene (PS) nanospheres. Human adipose-derived stem cell behaviors, such as cell morphology, adhesion, cytoskeleton contractility, proliferation, and differentiation, were investigated on the prepared PS nanopit surface. RESULTS: The osteogenic differentiation can be enhanced by nanopits with a diameter of 300-400 nm. DISCUSSION: The present study provided exciting new avenues to investigate cellular responses to well-defined nanoscale topographic features, which could further guide bone tissue engineering and stem cell clinical research. The capability to control developing biomaterials mimicking nanotopographic surfaces promoted functional tissue engineering, such as artificial joint replacement, bone repair, and dental applications.

High-density genetic map construction and identification of loci controlling flower-type traits in Chrysanthemum (Chrysanthemum × morifolium Ramat.).

Flower type is an important and extremely complicated trait of chrysanthemum. The corolla tube merged degree (CTMD) and the relative number of ray florets (RNRF) are the two key factors affecting chrysanthemum flower type. However, few reports have clarified the inheritance of these two complex traits, which limits directed breeding for flower-type improvement. In this study, 305 F1 hybrids were obtained from two parents with obvious differences in CTMD and RNRF performance. Using specific-locus amplified fragment sequencing (SLAF-seq) technology, we constructed a high-density genetic linkage map with an average map distance of 0.76 cM. Three major QTLs controlling CTMD and four major QTLs underlying RNRF were repeatedly detected in the 2 years. Moreover, the synteny between the genetic map and other Compositae species was investigated, and weak collinearity was observed. In QTL regions with a high degree of genomic collinearity, eight annotated genes were probed in the Helianthus annuus L. and Lactuca sativa L. var. ramosa Hort. genomes. Furthermore, 20 and 11 unigenes were identified via BLAST searches between the SNP markers of the QTL regions and the C. vestitum and C. lavandulifolium transcriptomes, respectively. These results lay a foundation for molecular marker-assisted breeding and candidate gene exploration in chrysanthemum without a reference assembly.

Image-guided cancer therapy using aptamer-functionalized cross-linked magnetic-responsive Fe3O4@carbon nanoparticles.

The excellent anticancer effect of combined differential cancer therapies has been observed in the last few decades. Efficient theragnostic nanoparticles (NPs) for malignancy treatment have received considerable research attention and widely investigated today. This study presents our results on the development of aptamer-functionalized Fe3O4@carbon@doxorubicin NPs (Apt-Fe3O4@C@DOX) and their application in the synergetic chemo-photothermal therapy (PTT) of cancer. The Apt-Fe3O4@C@DOX NPs displayed high photothermal conversion efficiency and extensive pH/heat-induced drug release. In vitro (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium) bromide experiments indicated that the combined chemo-PTT is much more toxic toward lung adenocarcinoma cells (A549) than PTT or chemotherapy alone. In addition, the Apt-Fe3O4@C@DOX NPs demonstrated decreasing contrast enhancement of magnetic resonance (MR) signals, which means they may be potentially applied as a contrast agent and serve as a critical component of T2-weighted MR imaging of tumor tissues. Taking the results together, the Apt-Fe3O4@C@DOX NPs show great potential for cancer therapy.

Reference Gene Selection for RT-qPCR Analysis of Flower Development in Chrysanthemum morifolium and Chrysanthemum lavandulifolium.

Quantitative real-time PCR (qPCR) is a popular and powerful tool used to understand the molecular mechanisms of flower development. However, the accuracy of this approach depends on the stability of reference genes. The capitulum of chrysanthemums is very special, which is consisting of ray florets and disc florets. There are obvious differences between the two types of florets in symmetry, gender, histological structure, and function. Furthermore, the ray florets have various shapes. The objective of present study was to identify the stable reference genes in Chrysanthemum morifolium and Chrysanthemum lavandulifolium during the process of flower development. In this study, nine candidate reference genes were selected and evaluated for their expression stability acrosssamples during the process of flower development, and their stability was validated by four different algorithms (Bestkeeper, NormFinder, GeNorm, and Ref-finder). SAND (SAND family protein) was found to be the most stably expressed gene in all samples or different tissues during the process of C. lavandulifolium development. Both SAND and PGK (phosphoglycerate kinase) performed most stable in Chinese large-flowered chrysanthemum cultivars, and PGK was the best in potted chrysanthemums. There were differences in best reference genes among varieties as the genetic background of them were complex. These studies provide guidance for selecting reference genes for analyzing the expression pattern of floral development genes in chrysanthemums.

Application of genomic SSR locus polymorphisms on the identification and classification of chrysanthemum cultivars in China.

The Chinese traditional chrysanthemum is a notable group of chrysanthemums (Chrysanthemum×morifolium Ramat.) in which the phenotypic characteristics richly vary. At present, there is a serious controversy regarding homonyms and synonyms within this group. Moreover, the current international chrysanthemum classification systems are not comprehensive enough to be used on Chinese traditional chrysanthemums. Thus, we first identified a broad collection of 480 Chinese traditional chrysanthemum cultivars using the unique DNA fingerprints and molecular identities that were established by 20 simple sequence repeat markers. Five loci, which distinguished all of the selected cultivars, were identified as the core loci to establish unique fingerprints and molecular identities with 19 denary digits for each cultivar. A cluster analysis based on Nei's genetic distance indicated that the selected cultivars were clustered according to their horticultural classification. Population structure analysis was subsequently performed with K values ranging from 2 to 14, and the most likely estimate for the population structure was ten subpopulations, which was nearly consistent with the clustering result. Principal component analysis was further performed to verify the classification results. On the basis of the Q-matrices of K = 10, a total of 19 traits were found to be associated with 42 markers. Taken together, these results can serve as starting points for the identification and classification of chrysanthemums based on the polymorphism of microsatellite markers, which is beneficial to promote the marker-assisted breeding and international communication of this marvelous crop.