Archimedean Spirals with Controllable Chirality: Disk Substrate-Mediated Solution Assembly of Rod-Coil Block Copolymers.

Spirals are common in nature; however, they are rarely observed in polymer self-assembly systems, and the formation mechanism is not well understood. Herein, we report the formation of two-dimensional (2D) spiral patterns via microdisk substrate-mediated solution self-assembly of polypeptide-based rod-coil block copolymers. The spiral pattern consists of multiple strands assembled from the block copolymers, and two central points are observed. The spirals fit well with the Archimedean spiral model, and their chirality is dependent on the chirality of the polypeptide blocks. As revealed by a combination of experiments and theoretical simulations, these spirals are induced by an interplay of the parallel ordering tendency of the strands and circular confinement of the microdisks. This work presents the first example regarding substrate-mediated self-assembly of block copolymers into spirals. The gained information could not only enhance our understanding of natural spirals but also assist in both the controllable preparations and applications of spiral nanostructures.

Living Growth Kinetics of Polymeric Micelles on a Substrate.

Living growth of micelles on the substrate is an intriguing phenomenon; however, little is known about its growth kinetics, especially from a theoretical viewpoint. Here, we examine the living growth kinetics of polymeric micelles on a hydrophobic substrate immersed in an aqueous solution. The block copolymers first assemble into short cylinder seeds anchored on the substrate. Then, the small aggregates of block copolymers in the solutions fuse onto the active ends of the anchored seeds, leading to micelle growth on the substrate. A theoretical model is proposed to interpret such living growth kinetics. It is revealed that the growth rate coefficient on the substrate is independent of the copolymer concentration and the multistep feedings; however, it is significantly affected by the surface hydrophobicity. Brownian dynamics simulations further support the proposed growth mechanism and the kinetic model. This work enriches living assembly systems and provides guidance for fabricating bioinspired surface nanostructures.

Highly parallel, wash-free, and ultrasensitive centrifugal droplet digital protein detection in sub-microliter blood.

The ability to efficiently detect low-abundance protein biomarkers in tiny blood samples is a significant challenge in clinical and laboratory settings. Currently, high-sensitivity approaches require specialized instrumentation, involve multiple washing steps, and lack the ability to parallelize, preventing their widespread implementation. Herein, we developed a parallelized, wash-free, and ultrasensitive centrifugal droplet digital protein detection (CDPro) technology that achieves a femtomolar limit of detection (LoD) of target proteins with sub-microliters of plasma. The CDPro combines two techniques, namely a centrifugal microdroplet generation device and a digital immuno-PCR assay. Miniaturized centrifugal devices enable emulsification of hundreds of samples within 3 minutes using a common centrifuge. The bead-free digital immuno-PCR assay not only eliminates the need for multistep washing, but also possesses ultra-high detection sensitivity and accuracy. We characterized the performance of CDPro using recombinant interleukins (IL-3 and IL-6) as example targets and reported a LoD of 0.0128 pg mL-1. We also quantified IL-6 from 7 human clinical blood samples using the CDPro with only 0.5 µL plasma, which showed excellent agreement with an existing clinical protein diagnostic system with 25 µL plasma from those samples (R2 = 0.98).

Comprehensive Analysis of Metabolome and Transcriptome in Fruits and Roots of Kiwifruit.

Kiwifruit (Actinidia chinensis) roots instead of fruits are widely used as Chinese medicine, but the functional metabolites remain unclear. In this study, we conducted comparative metabolome analysis between root and fruit in kiwifruit. A total of 410 metabolites were identified in the fruit and root tissues, and of them, 135 metabolites were annotated according to the Kyoto Encyclopaedia of Genes and Genomes (KEGG) pathway. Moreover, 54 differentially expressed metabolites (DEMs) were shared in root and fruit, with 17 DEMs involved in the flavonoid pathway. Of the 17 DEMs, three flavonols (kaempferol-3-rhamnoside, L-Epicatechin and trifolin) and one dihydrochalcone (phloretin) showed the highest differences in the content level, suggesting that flavonols and dihydrochalcones may act as functional components in kiwifruit root. Transcriptome analysis revealed that genes related to flavonols and dihydrochalcones were highly expressed in root. Moreover, two AP2 transcription factors (TFs), AcRAP2-4 and AcAP2-4, were highly expressed in root, while one bHLH TF AcbHLH62 showed extremely low expression in root. The expression profiles of these TFs were similar to those of the genes related to flavonols and dihydrochalcones, suggesting they are key candidate genes controlling the flavonoid accumulation in kiwifruit. Our results provided an insight into the functional metabolites and their regulatory mechanism in kiwifruit root.

Dynamic Transcriptome Analysis Reveals Complex Regulatory Pathway Underlying Induction and Dose Effect by Different Exogenous Auxin IAA and 2,4-D During in vitro Embryogenic Redifferentiation in Cotton.

Plant somatic cells can reprogram into differentiated embryos through somatic embryogenesis (SE) on the condition of plant growth regulators (PGRs). RNA sequencing analysis was performed to investigate transcriptional profiling on cotton redifferentiated callus that was induced by different auxin types (IAA and 2,4-D), different concentrations (0, 0.025, and 0.05 mg L-1), and different incubation times (0, 5, and 20 days). Under the 2,4-D induction effect, signal transduction pathways of plant hormones were significantly enriched in the embryogenic response stage (5 days). These results indicated that auxin signal transduction genes were necessary for the initial response of embryogenic differentiation. In the pre-embryonic initial period (20 days), the photosynthetic pathway was significantly enriched. Most differentially expressed genes (DEGs) were downregulated under the induction of 2,4-D. Upon the dose effect of IAA and 2,4-D, respectively, pathways were significantly enriched in phenylpropanoid biosynthesis, fatty acid metabolism, and carbon metabolic pathways. Therefore, primary and secondary metabolism pathways were critical in cotton SE. These results showed that complex synergistic mechanisms involving multiple cellular pathways were the causes of the induction and dose effect of auxin-induced SE. This study reveals a systematic molecular response to auxin signals and reveals the way that regulates embryogenic redifferentiation during cotton SE.

Anchorage-Dependent Living Supramolecular Self-Assembly of Polymeric Micelles.

Anchorage-dependent contact-inhibited growth usually refers to on-surface cell proliferation inhibited by the proximity of other cells. This phenomenon, prominent in nature, has yet to be achieved with polymeric micelles. Here, we report the control living supra-macromolecular self-assembly of elongated micelles with a liquid crystalline core onto a hydrophobic substrate via the synergetic interactions between the substrate and aggregates dispersed in solution. In this system, seed formation is a transient phenomenon induced by the adsorption and rearrangement of the core-swollen aggregates. The seeds then trigger the growth of elongated micelles onto the substrate in a living controllable manner until the contact with the substrate is disrupted. Brownian dynamic simulations show that this unique behavior is due to the fusion of the aggregates onto both ends of the anchored seeds. More important, the micelle length can be tuned by varying the substrate hydrophobicity, a key step toward the fabrication of intricate structures.

Quantitative metabolome and transcriptome analysis reveals complex regulatory pathway underlying photoinduced fiber color formation in cotton.

As an important plant single cell model and textile application materials, poorly known about fiber color formation in cotton, which is sensitively regulated by environmental signals. Our studies underline the importance of photo signal on sensitive fiber color formation and characterize fiber color early initiation (15 DPA) and late accumulated metabolites (45 DPA) in different lighting condition. The results revealed 236 differential metabolites between control and shading, of which phenylpropanoids metabolites accounted for 20%, including uncharacterized novel metabolites and pathways. Furthermore, the early initiation specific genes respond to the absence of light are highly correlated with phenylpropanoid metabolites related to pigmentation. The current study reveals the complex pathways involving early initiation regulation and late metabolic pathways. In addition, the collection composed of uncharacterized photoinduced metabolites and early initiation signaling/regulatory genes were identified, which are important resources for understanding fiber color formation. This report provides new insight into molecular regulatory and biochemical basis underlying photoinduced fiber color formation in cotton.

Quantitative analysis of low-concentration α-HMX based on terahertz spectroscopy.

Due to the instability of α type HMX at low concentrations, it belongs to the impurity crystal form. To ensure the functional effectiveness, operational reliability and management safety of HMX, it is necessary to quantify the low content of the unstable α-HMX crystal form in the composite explosive. In this study, low-concentration α-HMX is quantitatively analyzed in a mixture of α- and ß-HMX. First, terahertz time-domain spectroscopy (THz-TDS) is used to obtain the absorption spectrum of the α/ß-HMX element in the frequency range of 0.2-2.0 THz, and the characteristic frequency is selected. The absorption coefficient data in the frequency band of 0.7-1.3 THz are considered as the sample data for quantitative analysis. Finally, support vector machine (SVM) algorithm is used to establish a regression model, and principal component analysis (PCA) is employed for feature extraction. Grid search (GS), genetic algorithm (GA) and particle swarm optimization (PSO) are utilized for parameter optimization in support vector regression (SVR). These algorithms are combined to establish six regression models, and their effectiveness is assessed. The experimental results show that all the six methods can predict the content of α-HMX components with a small error and a high prediction accuracy. Compared to GA-SVR and PSO-SVR models, the PCA-GA-SVR and PCA-PSO-SVR models exhibit higher prediction accuracy and stability. The test set of the PCA-GA-SVR model reveals an average absolute error of 0.880%. It has the highest prediction accuracy, and the coefficient of determination (R2) reaches 0.9996. This indicates that PCA and SVR can be effectively used in the detection of low-concentration HMX components and can serve as a reliable basis for the quantitative analysis of other explosives.

2D Chiral Stripe Nanopatterns Self-Assembled from Rod-Coil Block Copolymers on Microstripes.

Chiral nanoarchitectures usually possess unique and intriguing properties. However, the construction of 2D chiral nanopatterns through polymer self-assembly is a challenge. Reported herein is the formation of chiral stripe nanopatterns through surface self-assembly of polypeptide-based rod-coil block copolymers on microstripes. The nanostripes align oblique to the boundary of the microstripes, resulting in the chirality of the nanopatterns. The chirality of the nanopatterns is closely related to the width of the microstripes, i.e., a narrower width results in higher chirality. Besides, the chiral sense of the nanopatterns can be regulated by the chirality of the polypeptide blocks. This work demonstrates the transmission of chirality from polymer to nanoarchitecture on a confined surface, which can guide the preparation of nanopatterns with tuned chiral features.

Adsorption and ordering of amphiphilic rod-coil block copolymers on a substrate: conditions for well-aligned stripe nanopatterns.

Controlling the ordering of self-assembled nanostructures is vital in block copolymer nanotechnology but still presents a challenge. Here we demonstrated that the adsorption and ordering of amphiphilic rod-coil block copolymers on a substrate can generate well-aligned stripe nanopatterns by dissipative particle dynamics simulations. The effects of the copolymer concentration and the surface affinity on the formation of stripe nanopatterns were examined. The simulation results revealed that the well-aligned stripe nanopatterns with controllable thickness and stripe width can be obtained in the systems with higher copolymer concentration and surface affinity. An immersion coating experiment was designed to verify the simulation results, and an agreement is shown. The present work provides a strategy for constructing well-aligned stripe nanopatterns in a controllable way.

Dynamic Transcriptome Analysis Reveals Uncharacterized Complex Regulatory Pathway Underlying Genotype-Recalcitrant Somatic Embryogenesis Transdifferentiation in Cotton.

As a notable illustration of totipotency and plant regeneration, somatic embryogenesis (SE) is the developmental reprogramming of somatic cells toward the embryogenesis pathway, the key step for genetic engineering. Investigations examining the totipotency process are of great fundamental and practical importance in crop biotechnology. However, high-frequency regeneration of cotton via SE has been limited due to genotype-dependent response. The molecular basis deciphering SE genotype recalcitrance remains largely unexplored in cotton. In the current study, to comprehensively investigate the dynamic transcriptional profiling and gene regulatory patterns involved in SE process, a genome-wide RNA sequencing analysis was performed in two cotton genotypes with distinct embryogenic abilities, the highly embryogenic genotype Yuzao 1 (YZ) and the recalcitrant genotype Lumian 1 (LM). Three typical developmental staged cultures of early SE-hypocotyls (HY), nonembryogenic calli (NEC) and primary embryogenic calli (PEC)-were selected to establish the transcriptional profiles. Our data revealed that a total of 62,562 transcripts were present amongst different developmental stages in the two genotypes. Of these, 18,394 and 26,514 differentially expressed genes (DEGs) were identified during callus dedifferentiation (NEC-VS-HY) and embryogenic transdifferentiation (PEC-VS-NEC), respectively in the recalcitrant genotype, 21,842 and 22,343 DEGs in the highly embryogenic genotype. Furthermore, DEGs were clustered into six expression patterns during cotton SE process in the two genotypes. Moreover, functional enrichment analysis revealed that DEGs were significantly enriched in fatty acid, tryptophan and pyruvate metabolism in the highly embryogenic genotype and in DNA conformation change otherwise in the recalcitrant genotype. In addition, critical SE-associated expressed transcription factors, as well as alternative splicing events, were notably and preferentially activated during embryogenic transdifferentiation in the highly embryogenic genotype compared with the recalcitrant genotype. Taken together, by systematically comparing two genotypes with distinct embryogenic abilities, the findings in our study revealed a comprehensive overview of the dynamic gene regulatory patterns and uncharacterized complex regulatory pathways during cotton SE genotype-dependent response. Our work provides insights into the molecular basis and important gene resources for understanding the underlying genotype recalcitrance during SE process and plant regeneration, thereby holding great promise for accelerating the application of biotechnology to cotton for improving its breeding efficiency.

Ethyl methanesulfonate mutant library construction in Gossypium hirsutum L. for allotetraploid functional genomics and germplasm innovation.

As the gene pool is exposed to both strain on land resources and a lack of diversity in elite allotetraploid cotton, the acquisition and identification of novel alleles has taken on epic importance in facilitating cotton genetic improvement and functional genomics research. Ethyl methanesulfonate (EMS) is an excellent mutagen that induces genome-wide efficient mutations to activate the mutagenic potential of plants with many advantages. The present study established, determined and verified the experimental procedure suitable for EMS-based mutant library construction as the general reference guide in allotetraploid upland cotton. This optimized method and procedure are efficient, and abundant EMS mutant libraries (approximately 12 000) in allotetraploid cotton were successfully obtained. More than 20 mutant phenotypes were observed and screened, including phenotypes of the leaf, flower, fruit, fiber and plant architecture. Through the plants mutant library, high-throughput and high-resolution melting technology-based variation evaluation detected the EMS-induced site mutation. Additionally, based on overall genome-wide mutation analyses by re-sequencing and mutant library assessment, the examination results demonstrated the ideal quality of the cotton EMS-treated mutant library constructed in this study with appropriate high mutation density and saturated genome. What is more, the collection is composed of a broad repertoire of mutants, which is the valuable resource for basic genetic research and functional genomics underlying complex allotetraploid traits, as well as cotton breeding.

Dynamic TMT-Based Quantitative Proteomics Analysis of Critical Initiation Process of Totipotency during Cotton Somatic Embryogenesis Transdifferentiation.

The somatic embryogenesis (SE) process of plants, as one of the typical responses to abiotic stresses with hormone, occurs through the dynamic expression of different proteins that constitute a complex regulatory network in biological activities and promotes plant totipotency. Plant SE includes two critical stages: primary embryogenic calli redifferentiation and somatic embryos development initiation, which leads to totipotency. The isobaric labels tandem mass tags (TMT) large-scale and quantitative proteomics technique was used to identify the dynamic protein expression changes in nonembryogenic calli (NEC), primary embryogenic calli (PEC) and globular embryos (GEs) of cotton. A total of 9369 proteins (6730 quantified) were identified; 805, 295 and 1242 differentially accumulated proteins (DAPs) were identified in PEC versus NEC, GEs versus PEC and GEs versus NEC, respectively. Eight hundred and five differentially abundant proteins were identified, 309 of which were upregulated and 496 down regulated in PEC compared with NEC. Of the 295 DAPs identified between GEs and PEC, 174 and 121 proteins were up- and down regulated, respectively. Of 1242 differentially abundant proteins, 584 and 658 proteins were up- and down regulated, respectively, in GEs versus NEC. We have also complemented the authenticity and accuracy of the proteomic analysis. Systematic analysis indicated that peroxidase, photosynthesis, environment stresses response processes, nitrogen metabolism, phytohormone response/signal transduction, transcription/posttranscription and modification were involved in somatic embryogenesis. The results generated in this study demonstrate a proteomic molecular basis and provide a valuable foundation for further investigation of the roles of DAPs in the process of SE transdifferentiation during cotton totipotency.

Metabolome and Transcriptome Association Analysis Reveals Dynamic Regulation of Purine Metabolism and Flavonoid Synthesis in Transdifferentiation during Somatic Embryogenesis in Cotton.

Plant regeneration via somatic embryogenesis (SE) is a key step during genetic engineering. In the current study, integrated widely targeted metabolomics and RNA sequencing were performed to investigate the dynamic metabolic and transcriptional profiling of cotton SE. Our data revealed that a total of 581 metabolites were present in nonembryogenic staged calli (NEC), primary embryogenic calli (PEC), and initiation staged globular embryos (GE). Of the differentially accumulated metabolites (DAMs), nucleotides, and lipids were specifically accumulated during embryogenic differentiation, whereas flavones and hydroxycinnamoyl derivatives were accumulated during somatic embryo development. Additionally, metabolites related to purine metabolism were significantly enriched in PEC vs. NEC, whereas in GE vs. PEC, DAMs were remarkably associated with flavonoid biosynthesis. An association analysis of the metabolome and transcriptome data indicated that purine metabolism and flavonoid biosynthesis were co-mapped based on the Kyoto encyclopedia of genes and genomes (KEGG) database. Moreover, purine metabolism-related genes associated with signal recognition, transcription, stress, and lipid binding were significantly upregulated. Moreover, several classic somatic embryogenesis (SE) genes were highly correlated with their corresponding metabolites that were involved in purine metabolism and flavonoid biosynthesis. The current study identified a series of potential metabolites and corresponding genes responsible for SE transdifferentiation, which provides a valuable foundation for a deeper understanding of the regulatory mechanisms underlying cell totipotency at the molecular and biochemical levels.