Effects of different light conditions on morphological, anatomical, photosynthetic and biochemical parameters of Cypripedium macranthos Sw.

In this study, the morphological (plant height, leaf length and width, stem diameter and leaf number), anatomical (epidermal cell density and thickness, Stomatal length and width), photosynthetic (net photosynthetic rate, transpiration rate, stomatal conductance, intercellular CO2 concentration, relative humidity, leaf temperature and chlorophyll fluorescence parameters) and biochemical parameters (the content of soluble sugar, soluble protein, proline, malondialdehyde and electrical conductivity) of Cypripedium macranthos Sw. in Changbai Mountain were determined under different light conditions (L10, L30, L50, L100). The results showed that morphological values including plant height, leaf area, stem diameter and leaf number of C. macranthos were smaller under the condition of full light at L100. The epidermal cell density and epidermal thickness of C. macranthos were the highest under L30 and L50 treatments, respectively. It had the highest net photosynthetic rate (Pn) and chlorophyll content under L50 treatment. Meanwhile, correlation analysis indicated that photosynthetically active radiation (PAR) and water use efficiency (WUE) were the main factors influencing Pn. C. macranthos accumulated more soluble sugars and soluble proteins under L100 treatment, while the degree of membrane peroxidation was the highest and the plant was severely damaged. In summary, the adaptability of C. macranthos to light conditions is ranked as follows L50 > L30 > L10 > L100. Appropriate light conditions for C. macranthos are 30%-50% of full light, which should be taken into account in protection and cultivation.

Time-Course Transcriptome Analysis of Aquilegia vulgaris Root Reveals the Cell Wall's Roles in Salinity Tolerance.

Salt stress has a considerable impact on the development and growth of plants. The soil is currently affected by salinisation, a problem that is becoming worse every year. This means that a significant amount of salt-tolerant plant material needs to be added. Aquilegia vulgaris has aesthetically pleasing leaves, unique flowers, and a remarkable tolerance to salt. In this study, RNA-seq technology was used to sequence and analyse the transcriptome of the root of Aquilegia vulgaris seedlings subjected to 200 mM NaCl treatment for 12, 24, and 48 h. In total, 12 Aquilegia vulgaris seedling root transcriptome libraries were constructed. At the three time points of salt treatment compared with the control, 3888, 1907, and 1479 differentially expressed genes (DEGs) were identified, respectively. Various families of transcription factors (TFs), mainly AP2, MYB, and bHLH, were identified and might be linked to salt tolerance. Gene Ontology (GO) analysis of DEGs revealed that the structure and composition of the cell wall and cytoskeleton may be crucial in the response to salt stress. Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis of the DEGs showed a significant enrichment of the pentose and glucuronate interconversion pathway, which is associated with cell wall metabolism after 24 and 48 h of salt treatment. Based on GO and KEGG analyses of DEGs, the pentose and glucuronate interconversion pathway was selected for further investigation. AP2, MYB, and bHLH were found to be correlated with the functional genes in this pathway based on a correlation network. This study provides the groundwork for understanding the key pathways and gene networks in response to salt stress, thereby providing a theoretical basis for improving salt tolerance in Aquilegia vulgaris.

Identification of Small RNAs Associated with Salt Stress in Chrysanthemums through High-Throughput Sequencing and Bioinformatics Analysis.

The Chrysanthemum variety "Niu 9717" exhibits excellent characteristics as an ornamental plant and has good salt resistance. In this study, this plant was treated with 200 mM NaCl for 12 h followed by high-throughput sequencing of miRNA and degradome. Subsequently, the regulatory patterns of potential miRNAs and their target genes were searched to elucidate how Chrysanthemum miRNAs respond to salt. From the root and leaf samples, we identified a total of 201 known miRNAs belonging to 40 families; furthermore, we identified 79 new miRNAs, of which 18 were significantly differentially expressed (p < 0.05). The expressed miRNAs, which targeted a total of 144 mRNAs in the leaf and 215 mRNAs in the root, formed 144 and 226 miRNA-target pairs in roots and leaves, respectively. Combined with the miRNA expression profile, degradome and transcriptome data were then analyzed to understand the possible effects of the miRNA target genes and their pathways on salt stress. The identified genes were mostly located in pathways related to hormone signaling during plant growth and development. Overall, these findings suggest that conserved and novel miRNAs may improve salt tolerance through the regulation of hormone signal synthesis or expression of genes involved in hormone synthesis.

Starch and Sucrose Metabolism and Plant Hormone Signaling Pathways Play Crucial Roles in Aquilegia Salt Stress Adaption.

Salt stress is one of the main abiotic stresses that strongly affects plant growth. Clarifying the molecular regulatory mechanism in ornamental plants under salt stress is of great significance for the ecological development of saline soil areas. Aquilegia vulgaris is a perennial with a high ornamental and commercial value. To narrow down the key responsive pathways and regulatory genes, we analyzed the transcriptome of A. vulgaris under a 200 mM NaCl treatment. A total of 5600 differentially expressed genes were identified. The Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis pointed out that starch and sucrose metabolism and plant hormone signal transduction were significantly improved. The above pathways played crucial roles when A. vulgaris was coping with salt stress, and their protein-protein interactions (PPIs) were predicted. This research provides new insights into the molecular regulatory mechanism, which could be the theoretical basis for screening candidate genes in Aquilegia.

Genome-wide identification and expression analysis of SBP-box gene family reveal their involvement in hormone response and abiotic stresses in Chrysanthemum nankingense.

SQUAMOSA promoter-binding-protein (SBP)-box family proteins are a class of plant-specific transcription factors, and widely regulate the development of floral and leaf morphology in plant growth and involve in environment and hormone signal response. In this study, we isolated and identified 21 non-redundant SBP-box genes in Chrysanthemum nankingense with bioinformatics analysis. Sequence alignments of 21 CnSBP proteins discovered a highly conserved SBP domain including two zinc finger-like structures and a nuclear localization signal region. According to the amino acid sequence alignments, 67 SBP-box genes from Arabidopsis thaliana, rice, Artemisia annua and C. nankingense were clustered into eight groups, and the motif and gene structure analysis also sustained this classification. The gene evolution analysis indicated the CnSBP genes experienced a duplication event about 10 million years ago (Mya), and the CnSBP and AtSPL genes occurred a divergence at 24 Mya. Transcriptome data provided valuable information for tissue-specific expression profiles of the CnSBPs, which highly expressed in floral tissues and differentially expressed in leaf, root and stem organs. Quantitative Real-time Polymerase Chain Reaction data showed expression patterns of the CnSBPs under exogenous hormone and abiotic stress treatments, separately abscisic acid, salicylic acid, gibberellin A3, methyl jasmonate and ethylene spraying as well as salt and drought stresses, indicating that the candidate CnSBP genes showed differentiated spatiotemporal expression patterns in response to hormone and abiotic stresses. Our study provides a systematic genome-wide analysis of the SBP-box gene family in C. nankingense. In general, it provides a fundamental theoretical basis that SBP-box genes may regulate the resistance of stress physiology in chrysanthemum via exogenous hormone pathways.

Chrysanthemum × grandiflora leaf and root transcript profiling in response to salinity stress.

As high soil salinity threatens the growth and development of plants, understanding the mechanism of plants' salt tolerance is critical. The Chrysanthemum × grandiflora is a newly developed species with a strong salt resistance that possesses multiple genes controlling its quantitative salt resistance. Because of this multigene control, we chose to investigate the plant stress genes overall responses at the transcriptome level. C. grandiflora were treated with a 200 mM NaCl solution for 12 h to study its effect on the roots and leaves via Illumina RNA sequencing. PAL, CYP73A, and 4CL in the phenylpropanoid biosynthesis pathway were upregulated in roots and leaves. In the salicylic acid signal transduction pathway, TGA7 was upregulated in the roots and leaves, while in the jasmonic acid signal transduction pathway, TIFY9 was upregulated in the roots and leaves. In the ion transporter gene, we identified HKT1 that showed identical expression patterns in the roots and leaves. The impact of NaCl imposition for 12 h was largely due to osmotic effect of salinity on C. grandiflora, and most likely the transcript abundance changes in this study were due to the osmotic effect. In order to verify the accuracy of the Illumina sequencing data, we selected 16 DEGs for transcription polymerase chain reaction (qRT-PCR) analysis. qRT-PCR and transcriptome sequencing analysis revealed that the transcriptome sequencing results were reliable.

An efficient approach for simultaneously obtaining oil and epigoitrin from Orychophragmus violaceus seeds by microwave-mediated immiscible binary solvent extraction.

Microwave-mediated immiscible binary solvent extraction (MIBSE) was applied to simultaneously extract oil and epigoitrin from Orychophragmus violaceus seeds. The upper phase of n-hexane was used to obtain oil, and the lower phase of ethanol solution was used to obtain epigoitrin. Factors potentially affecting the yields of oil and epigoitrin were systematically investigated. The optimum conditions were an ethanol volume fraction of 65%, liquid-solid ratio of lower-phase of 20 mL/g, liquid-solid ratio of upper-phase of 12 mL/g, microwave irradiation power of 393 W, and microwave irradiation time of 29 min. The actual yields of oil and epigoitrin were 34.08% ± 1.38% and 11.86 ± 0.47 mg/g, respectively. GC-MS analysis illustrated that the seed oils obtained by MIBSE and Soxhlet extraction exhibited similar fatty acid compositions. The separated epigoitrin was determined by HPLC analyses, which obtained a purity of 91.25% ± 3.83%, follwed by NMR determinations.

Syringa oblata genome provides new insights into molecular mechanism of flower color differences among individuals and biosynthesis of its flower volatiles.

Syringa oblata is a high ornamental value tree owing to its elegant colors, unique aromas and wide adaptability, however, studies on the molecular mechanism underlying the formation of its ornamental traits are still lacking. Here, we presented a chromosome-scale genome assembly of S. oblata and the final genome size was 1.11 Gb with a contig N50 of 4.75 Mb, anchored on 23 chromosomes and was a better reference for S. oblata transcriptome assembly. Further by integrating transcriptomic and metabolic data, it was concluded that F3H, F3'H, 4CL and PAL, especially the F3'H, were important candidates involved in the formation of floral color differences among S. oblata individuals. Genome-wide identification and analysis revealed that the TPS-b subfamily was the most abundant subfamily of TPS family in S. oblata, which together with the CYP76 family genes determined the formation of the major floral volatiles of S. oblata. Overall, our results provide an important reference for mechanistic studies on the main ornamental traits and molecular breeding in S. oblata.

Transgenic Chrysanthemum indicum overexpressing cin-miR396a exhibits altered plant development and reduced salt and drought tolerance.

The conserved microRNA396 (miR396) is involved in growth, development, and abiotic stress responses in a variety of plants by regulating target genes. Here, we obtained transgenic Chrysanthemum indicum (C. indicum) overexpressing the cin-miR396a gene. The transgenic plants (TGs) had longer internodes and fewer epidermal hairs in contrast with the wild-type (WT) control. cin-miR396a overexpression in C. indicum reduced salt tolerance and drought tolerance. After salt and drought stress compared with WT plants, the transgenic C. indicum exhibited a relative decrease in leaf water content, and the leaf free proline content, also exhibited a relative increase, in the leaf conductivity and leaf Malondialdehyde content, while the total chlorophyll content did not differ significantly from WT, and the Na+/K+ ratio in the roots of transgenic C. indicum increased after salt stress. We also identified two target genes of cin-miR396a, CiGRF1 and CiGRF5, whose expression was induced by salt and drought treatments and suppressed in transgenic C. indicum. Taken together, our results reveal a unique role for the regulatory module of miR396a-GRFs in C. indicum development and response to abiotic stresses. cin-miR396a plays a negative regulatory role in C. indicum in response to salt and drought stresses.

Overexpressing the NAC transcription factor LpNAC13 from Lilium pumilum in tobacco negatively regulates the drought response and positively regulates the salt response.

NACs are one of the largest transcription factor families in plants and are involved in the response to abiotic stress. A new stress-responsive NAC transcription factor gene, LpNAC13, was isolated from Lilium pumilum bulbs. The expression of LpNAC13 was induced by drought, salt, cold and ABA treatments. LpNAC13 overexpressing plants were generated to explore the function of LpNAC13 in response to drought and salt stress. Overexpression of LpNAC13 in tobacco displayed a reduced drought tolerance but exhibited an enhanced salt tolerance. The LpNAC13 overexpression plants had decreased antioxidant enzyme activities, content of proline and chlorophyll, increased MDA content under drought condition, the results in the LpNAC13 plants under salt condition were opposite to those under drought condition. The seed germination and root length assays of overexpression of LpNAC13 showed decreased sensitivity to ABA. Functional analyses demonstrate that LpNAC13 plays opposite roles in drought and salt stress tolerance, acting as a negative regulator of drought response but as a positive regulator of salt response in tobacco.

The complete chloroplast genome sequence of Tapiscia sinensis (Staphyleaceae).

Tapiscia sinensis, belong to Tapisciaceae, is endangered tree endemic to China. Here, we provide the complete plastid genomic data of T. sinensis with the aim of providing data for future conservation efforts research and revealing its phylogenetic position. The complete chloroplast sequence is 161,093 bp, including a large single-copy (LSC) region of 87,782 bp, a small single-copy (SSC) region of 18,517 bp, a pair of invert repeats (IR) regions of 27,387 bp. Plastid genome contains 131 genes, 85 protein-coding genes, 38 tRNA genes, and eight rRNA genes. Phylogenetic analysis base on 19 plastid genomes indicates that T. sinensis located Malvids branch, and is more closely related to the species of the order Sapindales than those of the order Malvales.

Transcriptome profiling provides insights into dormancy release during cold storage of Lilium pumilum.

BACKGROUND: Bulbs of the ornamental flower Lilium pumilum enter a period of dormancy after flowering in spring, and require exposure to cold for a period of time in order to release dormancy. Previous studies focused mainly on anatomical, physiological and biochemical changes during dormancy release. There are no dormancy studies of the northern cold-hardy wild species of Lilium at the molecular level. This study observed bulb cell and starch granule ultrastructures during cold storage; and analysed the transcriptome using sequencing. The combination of morphological and transcriptomic methods provides valuable insights into dormancy release during cold storage of Lilium pumilum. RESULTS: Ultrastructural changes reflected dormancy release during cold storage of the bulbs. We compared gene expression levels among samples at 0 (S1 stage), 30 (S2 stage), 60 (S3 stage) and 90 (S4 stage) d of cold storage, with 0 d as the control. The data showed that some regulatory pathways such as carbohydrate metabolism and plant hormone signal transduction were activated to break dormancy. Some differentially expressed genes (DEGs) related to antioxidant activity, epigenetic modification and transcription factors were induced to respond to low temperature conditions. These genes constituted a complex regulatory mechanism of dormancy release. CONCLUSIONS: Cytological data related to dormancy regulation was obtained through histomorphological observation; transcriptome sequencing provided comprehensive sequences and digital gene expression tag profiling (DGE) data, and bulb cell ultrastructural changes were closely related to DEGs. The novel Lilium pumilum genetic information from this study provides a reference for the regulation of dormancy by genetic engineering using molecular biology tools.

Overexpression of Chrysanthemum lavandulifolium ClCBF1 in Chrysanthemum morifolium 'White Snow' improves the level of salinity and drought tolerance.

This paper reports the first study on plant CBF transcription factors (TF) in salt and drought stress responses in Chrysanthemum lavandulifolium. A CBF homolog gene, named as ClCBF1, from C. lavandulifolium was isolated using rapid amplification of cDNA ends (RACE). The deduced peptide is comprised of 210 amino acids (AA) containing an AP2 structural domain characteristic of the AP2 gene family. Quantitative real-time PCR revealed that ClCBF1 gene exhibit differential expression patterns across root, leaf and stem tissues, and it was strongly induced under salt and drought treatments of C. lavandulifolium. Overexpression of ClCBF1 in C. morifolium 'White Snow' resulted in stronger tolerance to salt and drought stresses. The ClCBF1 expression level, enzymatic activities of superoxide dismutase and peroxidase, and contents of proline and soluble proteins were enhanced in these transgenic lines, they were repressed in the antisense transgenic lines under the same stress conditions. Results indicate that ClCBF1 represents a promising candidate gene in improving abiotic stress tolerance among ornamental plants.