Molecular Analysis of 14-3-3 Genes in Citrus sinensis and Their Responses to Different Stresses.

14-3-3 proteins (14-3-3s) are among the most important phosphorylated molecules playing crucial roles in regulating plant development and defense responses to environmental constraints. No report thus far has documented the gene family of 14-3-3s in Citrus sinensis and their roles in response to stresses. In this study, nine 14-3-3 genes, designated as CitGF14s (CitGF14a through CitGF14i) were identified from the latest C. sinensis genome. Phylogenetic analysis classified them into ε-like and non-ε groups, which were supported by gene structure analysis. The nine CitGF14s were located on five chromosomes, and none had duplication. Publicly available RNA-Seq raw data and microarray databases were mined for 14-3-3 expression profiles in different organs of citrus and in response to biotic and abiotic stresses. RT-qPCR was used for further examining spatial expression patterns of CitGF14s in citrus and their temporal expressions in one-year-old C. sinensis "Xuegan" plants after being exposed to different biotic and abiotic stresses. The nine CitGF14s were expressed in eight different organs with some isoforms displayed tissue-specific expression patterns. Six of the CitGF14s positively responded to citrus canker infection (Xanthomonas axonopodis pv. citri). The CitGF14s showed expressional divergence after phytohormone application and abiotic stress treatments, suggesting that 14-3-3 proteins are ubiquitous regulators in C. sinensis. Using the yeast two-hybrid assay, CitGF14a, b, c, d, g, and h were found to interact with CitGF14i proteins to form a heterodimer, while CitGF14i interacted with itself to form a homodimer. Further analysis of CitGF14s co-expression and potential interactors established a 14-3-3s protein interaction network. The established network identified 14-3-3 genes and several candidate clients which may play an important role in developmental regulation and stress responses in this important fruit crop. This is the first study of 14-3-3s in citrus, and the established network may help further investigation of the roles of 14-3-3s in response to abiotic and biotic constraints.

MicroRNAs and Transcripts Associated with an Early Ripening Mutant of Pomelo (Citrus grandis Osbeck).

'Liuyuezaoyou' is an early-ripening cultivar selected from a bud mutation of Citrus grandis Osbeck 'Guanximiyou'. They were designated here as MT and WT, respectively. The fruit of MT matures about 45 days earlier than WT, which was accompanied by significant changes in key phytohormones, sugar compounds and organic acids. Recent studies have showed that microRNAs (miRNAs) play an important role in regulation of fruit ripening process. The aim of this study was to compare MT fruits with WT ones to uncover if miRNAs were implicated in the ripening of C. grandis. Fruits of both WT and MT at four developmental stages were analyzed using high-throughput sequencing and RT-PCR. Several independent miRNA libraries were constructed and sequenced. A total of 747 known miRNAs were identified and 99 novel miRNAs were predicted across all libraries. The novel miRNAs were found to have hairpin structures and possess star sequences. These results showed that transcriptome and miRNAs are substantially involved in a complex and comprehensive network in regulation of fruit ripening of this species. Further analysis of the network model revealed intricate interactions of miRNAs with mRNAs during the fleshy fruit ripening process. Several identified miRNAs have potential targets. These include auxin-responsive protein IAA9, sucrose synthase 3, V-type proton ATPase, NCED1 (ABA biosynthesis) and PL1/5 (pectate lyase genes), as well as NAC100 putative coordinated regulation networks, whose interactions with respective miRNAs may contribute significantly to fruit ripening of C. grandis.

Volatiles Emitted at Different Flowering Stages of Jasminum sambac and Expression of Genes Related to α-Farnesene Biosynthesis.

Fresh jasmine flowers have been used to make jasmine teas in China, but there has been no complete information about volatile organic compound emissions in relation to flower developmental stages and no science-based knowledge about which floral stage should be used for the infusion. This study monitored volatile organic compounds emitted from living flowers of Jasminum sambac (L.) Ait. 'Bifoliatum' at five developmental stages and also from excised flowers. Among the compounds identified, α-farnesene, linalool, and benzyl acetate were most abundant. Since α-farnesene is synthesized through the Mevalonate pathway, four genes encoding 3-hydroxy-3-methylglutaryl coenzyme A synthase, 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGR), farnesyl pyrophosphate synthase, and terpene synthase were isolated. Their expression patterns in living flowers at the five stages and in excised flowers coincided with the emission patterns of α-farnesene. Application of lovastatin, a HMGR inhibitor, significantly reduced the expression of the genes and greatly decreased the emission of α-farnesene. The sweet scent was diminished from lovastatin-treated flowers as well. These results indicate that α-farnesene is an important compound emitted from jasmine flowers, and its emission patterns suggest that flowers at the opening stage or flower buds 8 h after excision should be used for the infusion of tea leaves.

Gear-Shaped High-g Combustion Chamber for Micro Turbojet Engine Applications.

Enhancing combustion efficiency and optimizing the thrust-to-weight ratio are critical technical challenges encountered in the development, application, and growth of micro turbojet engines. The high-centrifugal (high-g) combustion chamber, as an innovative combustion chamber system, has the capability to replace the primary combustion chamber of the traditional turbojet engine, reducing the length of the combustion chamber while maintaining engine performance. Previous studies on the structure of the high-g combustor (HGC) have shown problems such as uneven temperature distribution of the turbojet rotor. To improve the feasibility of HGC integration into micro turbojet engines, this study conducts relevant experiments on a 120 N thrust engine. Subsequently, the results of these experiments were used to analyze the structural design of HGC through a simulation approach. Including six main configurations, the first four structural designs focused on establishing a suitable highly centrifugal environment to stabilize and improve the combustion performance, which was successfully achieved by designing the outer ring gear-shaped inlet with four different angles. Subsequent structural designs were based around improving the uniformity of the temperature distribution at the combustion chamber outlet. The final design of the HGC combustion efficiency is not much different from the original combustion chamber, and it can shorten the axial length of the combustion chamber by nearly 30%. The design of the air inlet holes and the baffle plate effectively improves the temperature uniformity at the outlet of the combustion chamber. Moreover, without changing the combustion chamber material, the corresponding engine weight can be reduced by about 10.7%, and the engine thrust-to-weight ratio can be improved by up to 12% with the same thrust, which provides design ideas for further lightweight applications.

Prediction and Verification of Performance and Emission Characteristics of Diesel/Natural Gas Dual-Fuel Engine Based on Intelligent Algorithm.

With the development of computer application technologies, intelligent algorithm has been widely used in various fields. In this study, a coupled Gaussian process regression and feedback neural network (GPR-FNN) algorithm is proposed, and it is used to predict the performance and emission characteristics of a six-cylinder heavy-duty diesel/natural gas (NG) dual-fuel engine. Using the engine speed, torque, NG substitution rate, diesel injection pressure, and injection timing as inputs, an GPR-FNN model is established to predict the crank angle corresponding to 50% heat release, brake-specific fuel consumption, brake thermal efficiency, and carbon monoxide, carbon dioxide, total unburned hydrocarbon, nitrogen oxides, and soot emissions. Subsequently, its performance is evaluated using experimental results. The results show that the regression correlation coefficients of all output parameters are greater than 0.99, and the mean absolute percentage error is less than 5.9%. In addition, a contour plot is used to compare the experimental results with the GPR-FNN prediction data in detail, and the results show that the prediction model has high accuracy. The results of this study can provide new ideas for the research on diesel/natural gas dual-fuel engines.

Schiff base nanoarchitectonics for supramolecular assembly of dipeptide as drug carriers.

Development of peptide-based supramolecular materials with hierarchical morphology and tunable guest loading displays broad potential as drug carrier in view of biocompatibility and biodegradability. Herein, we report a facile Schiff base nanoarchitectonic for supramolecular assembly of diphenylalanine (FF) metastable gel. The addition of trace glutaraldehyde (GA)/H2O solution induces the Schiff base reaction between GA and FF accompanying by phase transition from gel to solution. FF nanoparticles and hierarchical beaded nanofibers with autofluorescence properties can be constructed by regulating the competitive assembly between FF-H2O and FF-GA oligomer. Moreover, various guest molecules with different hydrophilic and hydrophobic properties can be easily loaded into such assembled particles and its release can be triggered under weak alkaline conditions, which show the potential application of the assembled FF system as drug carriers.

[Transcriptome analysis of flowering regulated by red and blue light in Jasminum sambac].

Jasminum sambac is a perennial evergreen shrub plant. With fragrant and aroma, and often used as a raw material for natural spices. In this study, we used white light as the control group, red-light and blue-light as the treatment to study effects of different light on jasmine flowering. Red- light promoted jasmine flowering in advance and increased the number of flower buds, whereas blue-light delayed jasmine flowering and decreased the number of flower buds. There was significant difference on the number of flower buds among the three groups. The top buds' transcriptomes of different light were sequenced by the Illumina Hiseq/Miseq 2000 high-throughput sequencing technology. In total 2 452 457 Unigenes were generated by transcriptome sequencing, of which 1 760 723 Unigenes were annotated into GO, COG, KEGG, KOG, NR, Pfam, Swiss-Prot, NOG databases. There were 894 DEGs in the control group vs red-light group, 2 690 DEGs in the control group vs blue-light group, and 3 828 DEGs in the red-light group vs blue-light group. KEGG Enrichment analysis reveals that the significant enrichment pathways had 6 pathways, including secondary metabolite biosynthesis, phenylpropanoid biosynthesis, indole alkaloid biosynthesis, photosynthesis, plant hormone signaling, and plants-pathogen interactions, and 24 related DEGs were detected by RT-qPCR, the result of which was significantly correlated with the transcriptome data. Through further analysis of transcriptome data, a large number of flowering-related hormones (IAA, ETH, GA, CTK, ABA, SA, JA) signal transduction genes and flowering pathway-related regulatory genes (PHY, CRY1, FPA, AGL and SOC1) and transcription factor (bHLH, MYB, WKRY) family genes were found. The study will help elucidate the differential expression mechanism of different light regulation of jasmine flowering.

CuS@Corn Stalk/Chitin Composite Hydrogel for Photodegradation and Antibacterial.

Copper sulfide nanoparticles (CuS NPs) have recently attracted extensive attention in various fields due to their excellent optical and electrical properties. However, CuS NPs are easy to agglomerate in their preparation on account of the high surface activity. In this study, uniform dispersion of CuS NPs were fabricated with corn stalk as a template and stabilizer, further CuS@corn stalk/chitin composite hydrogel was obtained by crosslinking with chitin. The results reveal that the CuS NPs were evenly dispersed into the composite hydrogels with a three-dimensional network structure, which were verified by the UV-vis spectrum, XRD, FT-IR spectra and SEM. In addition, the as-prepared composite hydrogel with the traits of peroxidase-like activity can convert H2O2 into an extremely oxidative and toxic ·OH, which manifested good effects for photodegradation of RhB and antibacterial against Escherichia coli and Staphylococcus aureus. Hence, the composite hydrogels could be used for photocatalytic treatment and sterilization of wastewater, which provides a new idea for the functional application of CuS NPs.

Identification of Appropriate Reference Genes for Normalizing miRNA Expression in Citrus Infected by Xanthomonas citri subsp. citri.

MicroRNAs (miRNAs) are short noncoding RNA molecules that regulate gene expression at the posttranscriptional level. Reverse transcription-quantitative PCR (RT-qPCR) is one of the most common methods used for quantification of miRNA expression, and the levels of expression are normalized by comparing with reference genes. Thus, the selection of reference genes is critically important for accurate quantification. The present study was intended to identify appropriate miRNA reference genes for normalizing the level of miRNA expression in Citrus sinensis L. Osbeck and Citrus reticulata Blanco infected by Xanthomonas citri subsp. citri, which caused citrus canker disease. Five algorithms (Delta Ct, geNorm, NormFinder, BestKeeper and RefFinder) were used for screening reference genes, and two quantification approaches, poly(A) extension RT-qPCR and stem-loop RT-qPCR, were used to determine the most appropriate method for detecting expression patterns of miRNA. An overall comprehensive ranking output derived from the multi-algorithms showed that poly(A)-tailed miR162-3p/miR472 were the best reference gene combination for miRNA RT-qPCR normalization in citrus canker research. Candidate reference gene expression profiles determined by poly(A) RT-qPCR were more consistent in the two citrus species. To the best of our knowledge, this is the first systematic comparison of two miRNA quantification methods for evaluating reference genes. These results highlight the importance of rigorously assessing candidate reference genes and clarify some contradictory results in miRNA research on citrus.