Utilizing transcriptomics and metabolomics to unravel key genes and metabolites of maize seedlings in response to drought stress.

BACKGROUND: Drought stress can substantially restrict maize growth and productivity, and global warming and an increasing frequency of extreme weather events are likely to result in more yield losses in the future. Therefore, unraveling the molecular mechanism underlying the response to drought stress is essential for breeding drought-resilient crops. RESULTS: In this study, we subjected the 3-leaf-period plants of two maize inbred lines, a drought-tolerant line (si287) and a drought-sensitive line (X178), to drought stress for seven days while growing in a chamber. Subsequently, we measured physiological traits and analyzed transcriptomic and metabolic profiles of two inbred lines. Our KEGG analysis of genes and metabolites revealed significant differences in pathways related to glycolysis/gluconeogenesis, flavonoid biosynthesis, starch and sucrose metabolism, and biosynthesis of amino acids. Additionally, our joint analysis identified proline, tryptophan and phenylalanine are crucial amino acids for maize response to drought stress. Furthermore, we concentrated on tryptophan (Trp), which was found to enhance tolerance via IAA-ABA signaling, as well as SA and nicotinamide adenine dinucleotide (NAD) consequent reactive oxygen species (ROS) scavenging. We identified three hub genes in tryptophan biosynthesis, indole-3-acetaldehyde oxidase (ZmAO1, 542,228), catalase 1 (ZmCAT1, 542,369), and flavin-containing monooxygenase 6 (ZmYUC6, 103,629,142), High expression of these genes plays a significant role in regulating drought tolerance. Two metabolites related to tryptophan biosynthesis, quinolinic acid, and kynurenine improved maize tolerance to drought stress by scavenging reactive oxygen species. CONCLUSIONS: This study illuminates the mechanisms underlying the response of maize seedlings to drought stress. Especially, it identifies novel candidate genes and metabolites, enriching our understanding of the role of tryptophan in drought stress. The identification of distinct resistance mechanisms in maize inbred lines will facilitate the exploration of maize germplasm and the breeding of drought-resilient hybrids.

A P-type pentatricopeptide repeat protein ZmRF5 promotes 5' region partial cleavages of atp6c transcripts to restore the fertility of CMS-C maize by recruiting a splicing factor.

A fast evolution within mitochondria genome(s) often generates discords between nuclear and mitochondria, which is manifested as cytoplasmic male sterility (CMS) and fertility restoration (Rf) system. The maize CMS-C trait is regulated by the chimeric mitochondrial gene, atp6c, and can be recovered by the restorer gene ZmRf5. Through positional cloning in this study, we identified the nuclear restorer gene, ZmRf5, which encodes a P-type pentatricopeptide repeat (PPR) family protein. The over-expression of ZmRf5 brought back the fertility to CMS-C plants, whereas its genomic editing by CRISPR/Cas9 induced abortive pollens in the restorer line. ZmRF5 is sorted to mitochondria, and recruited RS31A, a splicing factor, through MORF8 to form a cleaving/restoring complex, which promoted the cleaving of the CMS-associated transcripts atp6c by shifting the major cleavage site from 480th nt to 344 th nt for fast degradation, and preserved just right amount of atp6c RNA for protein translation, providing adequate ATP6C to assembly complex V, thus restoring male fertility. Interestingly, ATP6C in the sterile line CMo17A, with similar cytology and physiology changes to YU87-1A, was accumulated much less than it in NMo17B, exhibiting a contrary trend in the YU87-1 nuclear genome previously reported, and was restored to normal level in the presence of ZmRF5. Collectively these findings unveil a new molecular mechanism underlying fertility restoration by which ZmRF5 cooperates with MORF8 and RS31A to restore CMS-C fertility in maize, complemented and perfected the sterility mechanism, and enrich the perspectives on communications between nucleus and mitochondria.

Split G-quadruplex based PfAgo sensing platform for nucleotide mutation discrimination and human genotyping.

A PfAgo-G4 sensing platform exploiting G4 as a signal reporter was proposed, validated, and optimized. By introducing two mismatches at the Link strand, a universal nucleotide design rule was established for accurate single nucleotide polymorphism discrimination with PfAgo-G4. The FUT2 gene was then successfully and accurately genotyped using human buccal swab samples.

First Report of Leaf Spot Disease on Lonicera japonica Caused by Nigrospora oryzae in China.

Lonicera japonica Thunb. is a traditional Chinese medicinal plant, which widely cultivated in China, Japan and Korea. From August to October in 2021 and 2022, severe leaf spots symptoms were observed on L. japonica in medicinal botanical garden of Shandong University of Traditional Chinese Medicine (36°55'89"N, 116°79'91"E), Jinan, Shandong Province, China. The disease incidence was above 80% in the 25 acre cultivation area. Early symptoms were small brown spots on the leaves. Then the number of small spots gradually increased and spread over the entire leaves. The small brown spots seldom merge together to form larger lesions. Leaves with typical symptoms were collected from twenty individual plants, and cut into small 5×5 mm fragments in the junction of infected and healthy tissues. The fragments were sterilized in 75% ethanol for 30 s and 1% NaClO for 60 s, rinsed three times in sterile water, and then placed on potato dextrose agar (PDA). After 3 days of incubation at 25°C, fungal plugs along the edge of the colony were cut and transferred to new PDA for purification. A total number of 23 colonies with similar morphological characteristics were obtained, and three representative strains (Lj14, Lj18 and Lj20) were selected for subsequent study. The colonies grew rapidly on PDA and covered the entire petri dish in 4 days. Colonies had abundant aerial hyphae, initially white, round, later turning gray and black. Conidia were oblate or nearly spherical, single-celled, black, and measured in size from 9.6 to 13.2 µm × 7.9 to 16.1 µm in diameter (n=150) (Figure S1). The observed characteristics were close to those of Nigrospora spp. ( Wang et al. 2017). The genomic DNA was extracted, and PCR amplification of the rDNA internal transcribed spacer (ITS), ß-tubulin gene (TUB), and translation elongation factor 1-alpha gene (TEF1) were completed by primers ITS1/ITS4, Bt2a/Bt2b and EF1-728F/EF1-986R (Carbone and Kohn, 1999). Sequences were deposited in GenBank (accession nos. OR936661, OR936662, OR936671 for ITS, OR947626, OR947627, OR947628 for TUB, and OR947629, OR947630, OR947631 for TEF1 sequences, respectively). BLAST analyses of ITS (OR936661), TUB(OR947626) and TEF1 (OR947629) sequences exhibited 100% (487 bp out of 487 bp), 99.48% (380 bp out of 382 bp), and 99.6% (248 bp out of 249 bp) similarity to the sequences of N. oryzae strains KoLRI_053384 (MZ855426), LC2991 (KY019496) and LC7307 (KY019409), respectively. Lj14, Lj18 and Lj20 formed a clade with N. oryzae LC6763 and LC2991 in phylogenetic tree (Figure S2). Based on morphological and molecular evidence, the pathogen was identified as N. oryzae (Berk. &Broome) Petch. To fulfill Koch's postulates, the pathogenicity was tested in vivo experiments. Thirty non-wounded healthy leaves of ten intact plants were inoculated with 10 µl spore suspension (106 spores/ml) of three strains, respectively. As negative control, thirty leaves of ten healthy plants were inoculated with sterile water. The inoculated plants were placed at 28°C in the growth chamber with high relative humidity. The pathogenicity tests were repeated three times. Distinct symptoms similar to that of natural conditions were observed on the leaves of inoculated plants after 4 to 7 days. The strain was reisolated from the lesions and identified as N. oryzae by morphological features and ITS sequence. The pathogen has been reported to cause leaf spot disease on tobacco (Wang et al. 2022) and asiatic dayflower (Qiu et al. 2022). To our knowledge, this is the first report of leaf spot caused by N. oryzae on Lonicera japonica in China. The research will be helpful for leaf spot disease control.

Diversity and functional prediction of fungal communities in different segments of mongolian horse gastrointestinal tracts.

BACKGROUND: Anaerobic fungi are effective fibre-degrading microorganisms in the digestive tract of horses. However, our understanding of their diversity and community structure is limited, especially in different parts of the gastrointestinal tract. RESULTS: For the first time, high-throughput sequencing technology was used to analyse and predict fungal microbial diversity in different parts of the gastrointestinal tract of Mongolian horses. The results revealed that the richness and diversity of fungi in the hindgut of Mongolian horses were much higher than those in the foregut. The foregut was dominated by Basidiomycota and Ascomycota, whereas the hindgut was dominated by Neocallimastigomycota and Basidiomycota. At the genus level, the relative abundance of many pathogenic fungi (Cryptococcus, Cladosporium, Alternaria, and Sarocladium) in the foregut was significantly higher than that in the posterior gut, indicating that Mongolian horses have strong disease resistance. The prediction of fungal function also showed significant differences in the fungal flora between the foregut and the hindgut. The fungi in Mongolian horses' foreguts were mainly pathologically nutritive and contained many animal and plant pathogens, particularly in the small intestine (jejunum and ileum). This indicates that the foregut may be the most important immune site in the digestive system of Mongolian horses, which explains the high disease resistance of Mongolian horses. The number of unassigned functional groups in the posterior gut was significantly higher than that in the anterior gut, indicating that the functions of fungal groups in the posterior gut have not been fully explored, and further studies are required in the future. CONCLUSIONS: Analysis of high-throughput sequencing results revealed that the fungal composition varied greatly among different gastrointestinal tract segments in Mongolian horses, whose hindgut contains many anaerobic fungi involved in plant cellulose degradation. This provides important basic data for studying fungal diversity in the digestive system of healthy horses, which can be used for the health assessment of horses and provides clues for further research on the disease resistance and digestive capacity of horses, as well as a reference for the early diagnosis of intestinal diseases and innovative treatment methods.

Rice Cell Division Cycle 20s are required for faithful chromosome segregation and cytokinesis during meiosis.

Chromosome segregation must be under strict regulation to maintain chromosome euploidy and stability. Cell Division Cycle 20 (CDC20) is an essential cell cycle regulator that promotes the metaphase-to-anaphase transition and functions in the spindle assembly checkpoint, a surveillance pathway that ensures the fidelity of chromosome segregation. Plant CDC20 genes are present in multiple copies, and whether CDC20s have the same functions in plants as in yeast and animals is unclear, given the potential for divergence or redundancy among the multiple copies. Here, we studied all three CDC20 genes in rice (Oryza sativa) and constructed two triple mutants by clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein 9-mediated genome editing to explore their roles in development. Knocking out all three CDC20 genes led to total sterility but did not affect vegetative development. Loss of the three CDC20 proteins did not alter mitotic division but severely disrupted meiosis as a result of asynchronous and unequal chromosome segregation, chromosome lagging, and premature separation of chromatids. Immunofluorescence of tubulin revealed malformed meiotic spindles in microsporocytes of the triple mutants. Furthermore, cytokinesis of meiosis I was absent or abnormal, and cytokinesis II was completely prevented in all mutant microsporocytes; thus, no tetrads or pollen formed in either cdc20 triple mutant. Finally, the subcellular structures and functions of the tapetum were disturbed by the lack of CDC20 proteins. These findings demonstrate that the three rice CDC20s play redundant roles but are indispensable for faithful meiotic chromosome segregation and cytokinesis, which are required for the production of fertile microspores.

Preoperative cerebrospinal fluid biomarkers may be associated with postoperative delirium in patients undergoing knee/hip arthroplasty: the PNDABLE study.

BACKGROUND: In the global aging population, the incidence of postoperative delirium (POD) is increasing. Therefore, finding its effective predictive tools becomes crucial. We aimed to identify potential Cerebrospinal fluid (CSF)biomarkers for POD. METHODS: A total of 825 patients undergoing knee/hip arthroplasty under combined spinal-epidural anesthesia were selected. The patients were aged 40 to 90 years with American Society of Anesthesiologists physical status I~II. The Mini-Mental State Examination was completed 1 day before the operation. CSF was extracted after successful spinal-epidural combined puncture, and α-synuclein (α-syn), amyloid beta40 (Aß40), amyloid beta42 (Aß42), t-Tau, phosphorylated Tau (p-Tau), progranulin (PGRN) and soluble triggering receptor expressed on myeloid cells 2 (sTREM2) in the CSF were measured by enzyme-linked immunosorbent assays (ELISA). The patient's operation time, anesthesia time, intraoperative blood loss and fluid input were also recorded. After the operation, the occurrence rate and severity of POD were determined by the Confusion Assessment Method and the Memorial Delirium Assessment Scale (MDAS), respectively. Patients were categorized into POD group and non-POD group. Logistic regression analysis was performed on the indicators with statistically significant differences, and the area under the ROC curve (AUC) was used to estimate the predictive accuracy of the biomarkers for POD. RESULTS: A total of 92 patients developed POD and the incidence of POD was 11.15%. The results of the multivariable logistic regression showed that CSF t-Tau (P = 0.004, OR = 1.006, 95%CI 1.002~1.009) and α-syn (P = 0.004, OR = 1.001, 95%CI 1.000~1.001) were positively associated with the occurrence rate of POD, while Aß42 (P < 0.001, OR = 0.989, 95%CI 0.986~0.993), CSF PGRN (P = 0.002, OR = 0.999, 95%CI 0.999~1.000), Aß42/ t-Tau (P < 0.001, OR = 0.181, 95%CI 0.102~0.319) and Aß42/p-Tau (P < 0.001, OR = 0.617, 95%CI 0.526~0.725) were inversely proportional to the occurrence of POD. ROC curve analysis indicated that Aß42/t-Tau (AUC = 0.823), CSF Aß42 (AUC = 0.813), Aß42/p-Tau (AUC = 0.810), α-syn (AUC = 0.644) and PGRN (AUC = 0.638) could predict the occurrence rate of POD. The combination of all these biomarkers showed a greater AUC(0.896) than using any of them alone. CONCLUSIONS: CSF Aß42, PGRN, α-syn, Aß42/t-Tau and Aß42/p-Tau might be associated with the occurrence rate of POD in patients undergoing knee/hip arthroplasty. TRIAL REGISTRATION: Clinical Registration No. ChiCTR2000033439.

Coacervate Formed by an ATP-Binding Aptamer and Its Dynamic Behavior under Nonequilibrium Conditions.

Although numerous protocell models have been developed to explore the possible pathway of the origin of life on the early earth, few truly fulfill the roles of the DNA/RNA sequence and ATP molecules, which are keys to cell replication and evolution. The ATP-binding aptamer offers an opportunity to combine sequence and energy molecules. In this work, we choose the coacervate droplet as the protocell model and investigate the interaction of the DNA aptamer, poly(l-lysine)(PLL), and ATP under varying conditions. PLL and aptamers form solid precipitates, which spontaneously transform to coacervate droplets as ATP is introduced. The selective uptake and sequestration of exogenous molecules is achieved by the ATP-containing coacervates. As an electric field is applied to expel ATP, the portion of the droplet deficient in ATP becomes solid. The solid/liquid phase ratio is tunable by varying the electric field strength and excitation time. Together with the vacuolization process, a solid head with a soft mouth periodically opening and closing is created. Moreover, the composite coacervate droplet gradually grows as it is treated with an electric field and cannot recover to the original liquid phase after the power is turned off and replenished with ATP. Our work highlights that the proper integration of the DNA sequence, ATP, and energy input could be a powerful strategy for creating a protocell with certain living features.

The effect of neostigmine on postoperative delirium after colon carcinoma surgery: a randomized, double-blind, controlled trial.

BACKGROUND: Postoperative delirium (POD) is a critical complication in patients accepting colon carcinoma surgery. Neostigmine, as a cholinesterase inhibitor, can enhance the transmission of cholinergic transmitters in synaptic space, and play an important role in maintaining the normal level of cognition, attention and consciousness. The objective of this study was to investigate the effect of neostigmine on POD and clinical prognosis. METHODS: A randomized, double-blind controlled trial was implemented in Qingdao Municipal Hospital Affiliated to Qingdao University. A total of 454 patients aged 40 to 90 years old accepted colon carcinoma surgery were enrolled between June 7, 2020, and June 7, 2021, with final follow-up on December 8, 2021. Patients were randomly assigned to two groups: the neostigmine group (group N) and the placebo group (group P), the patients in group N were injected with 0.04 mg/kg neostigmine and 0.02 mg/kg atropine intravenously. The primary endpoint was the incidence of POD, researchers evaluated the occurrence of POD by the Confusion Assessment Method (CAM) twice daily (at 10 a.m. and 2 p.m.) during the first 7 postoperative days, POD severity was assessed by the Memorial Delirium Assessment Scale (MDAS). The secondary endpoints were the extubating time, postanesthesia care unit (PACU) time, the incidence of various postoperative complications, length of hospital stays, and 6 months postoperative mortality. RESULTS: The incidence of POD was 20.20% (81/401), including 19.39% (38/196) in group N and 20.98% (43/205) in group P. There was no significant statistical significance in the incidence of POD between group N and group P (P > 0.05); Compared to group P, the extubating time and PACU time in group N were significantly reduced (P < 0.001), the incidence of postoperative pulmonary complications (POPCs) decreased significantly in group N (P < 0.05), while no significant differences were observed in postoperative hospital stay and mortality in 6 months between the two groups (P > 0.05). CONCLUSION: For patients accepted colon carcinoma surgery, neostigmine did not significantly reduce the incidence of POD, postoperative mortality and postoperative hospital stay, while it indeed reduced the extubating time, PACU time and the incidence of POPCs. TRIAL REGISTRATION: The randomized, double-blind, controlled trial was registered retrospectively at www.chictr.org.cn on 07/06/2020 (ChiCTR2000033639).

Correlation between microRNA-320 and postoperative delirium in patients undergoing tibial fracture internal fixation surgery.

BACKGROUND: Although the incidence of postoperative delirium (POD) in the elderly after surgery are rising as individuals are living longer, the pathogenesis of POD remains poorly understood. It has been suggested that miRNA-320 may play a role in POD based on animal study and human study. METHODS: We first carried out an animal study, and designed and conducted a human study based on the result of animal study. The aged rats were randomly assigned to five groups: the control (C), anesthesia and surgery (AS), saline (NS), agomir-320 (AG), and antagomir-320 (AT) groups. Postoperative spatial learning and memory in rats were analyzed by the Morris water maze and the open field tests. The plasma levels of insulin-like growth factor-1 (IGF-1), amyloid precursor protein (APP) proteins, miRNA320 and IGF-1mRNA were measured by ELISA and qRT-PCR, respectively. A total of 240 Chinese Han patients over 65 years who underwent tibial fracture internal fixation were included in the PNDABLE study. POD cases and non-POD controls (1:1 matched) were selected by an anesthesiologist using Confusion Assessment Method. RESULTS: For Group AS, the escape latency was significantly longer and the ratio of time spent in the target quadrant was significantly reduced, APP and miR-320 were upregulated and IGF-1mRNA was downregulated compared with Group C. For Group AG, the escape latency was significantly longer and the ratio of time spent in the target quadrant was significantly reduced, APP and miR-320 were upregulated and IGF-1mRNA was downregulated compared with Group AS. For Group AT, the escape latency was significantly reduced and the ratio of time spent in the target quadrant was significantly longer, APP and miR-320 were downregulated and IGF-1mRNAwas upregulated compared with Group AS. Compared with NPOD patients, the expressions of plasma miR-320 and APP protein were increased and the expression of plasma IGF-1 mRNA was decreased in POD patients after surgery. CONCLUSIONS: MiRNA-320 might play a role in up-regulating the levels of IGF-1mRNA and APP protein, which offered a new target for POD treatment. TRIAL REGISTRATION: Correlation of perioperative neurocognitive disorders with lifestyle and biomarkers. ChiCTR2000033439 . Registered 1 June 2020.

The nutrient requirements of Lactobacillus acidophilus LA-5 and their application to fermented milk.

Lactobacillus acidophilus LA-5 is a suitable probiotic for food application, but because of its slow growth in milk, an increase in its efficiency is desired. To shorten the time required for fermentation, the nutrient requirements of L. acidophilus LA-5 were analyzed, including the patterns of consumption of amino acids, purines, pyrimidines, vitamins, and metal ions. The nutrients required by L. acidophilus LA-5 were Asn, Asp, Cys, Leu, Met, riboflavin, guanine, uracil, and Mn2+, and when they were added to milk, the fermentation time of fermented milk prepared by L. acidophilus LA-5 alone was shortened by 9 h, with high viable cell counts that were maintained during storage of nutrient-supplemented fermented milk compared with the control. For fermented milk prepared by fermentation with Streptococcus thermophilus, Lactobacillus delbrueckii ssp. bulgaricus, and L. acidophilus LA-5, viable cell counts of L. acidophilus LA-5 increased 1.3-fold and were maintained during storage of nutrient-supplemented fermented milk compared with the control. Adding nutrients had no negative effect on the quality of the fermented milk. The results indicated that suitable nutrients enhanced the growth of L. acidophilus LA-5 and increased its viable cell counts in fermented milk prepared by L. acidophilus LA-5 alone and mixed starter culture, respectively.

Synthesis of Novel Spiro-Tetrahydroquinoline Derivatives and Evaluation of Their Pharmacological Effects on Wound Healing.

A highly diastereoselective method for the synthesis of novel spiro-tetrahydroquinoline derivatives is reported here, using a one-pot reaction method. All compounds were characterized by 1H-nuclear magnetic resonance (NMR) and mass spectroscopy, and their stereo configurations were confirmed by X-ray analysis. These activities of these derivatives were then tested in human keratocyte cells. The responses of cells to treatment with selected compounds were studied using scratch analysis, and the compounds were tested in a mouse excision wound model. Three of the derivatives demonstrated significant wound-healing activities.

Dynamic Behavior of Complex Coacervates with Internal Lipid Vesicles under Nonequilibrium Conditions.

During the evolution of life on earth, the emergence of lipid membrane-bounded compartments is one of the most enigmatic events. Endosymbiosis has been hypothesized as one of the solutions. In this work, using a coacervate droplet formed by single-stranded oligonucleotides (ss-oligo) and poly(l-lysine) (PLL) as the protocell model, we monitored the uptake of liposomes of different types and studied the dynamic behavior of the resulting composite droplet under the electric field. The coacervate droplet exhibits affinity for the liposomes of varying charges. However, the permeation of liposome is also controlled by electrostatic interactions. Dominated by electrostatic attraction, the positively charged liposome is retained inside the droplet as growing fibrous structures, while the negatively charged liposome is mainly coated on the droplet surface. Permeation and even distribution occur when the liposome and the droplet carry the same charges, or at least one of them is neutral. As an electric field is applied to trigger repetitive cycles of vacuolization in the ss-oligo/PLL droplet, the fibrous structure formed by the positively charged liposome is basically intact, while a new phase is generated together with uneven mass transport as the negatively charged liposome is internalized. Interestingly, the release of daughter droplets with similar components occurs on the droplet containing neutral liposomes. Our work not only provides a step toward creating protocells with hierarchical structures and biofunctions using a biogenetic material via simple mixing but also sheds light on the possible origin of the lipid structure inside a living organism.

Fission and Internal Fusion of Protocell with Membraneless "Organelles" Formed by Liquid-Liquid Phase Separation.

Construction of protocells with hierarchical structures and living functions is still a great challenge. Growing evidence demonstrates that the membraneless organelles, which facilitate many essential cellular processes, are formed by RNA, protein, and other biopolymers via liquid-liquid phase separation (LLPS). The formation of the protocell in the early days of Earth could follow the same principle. In this work, we develop a novel coacervate-based protocell containing membraneless subcompartments via spontaneous liquid-liquid phase separation by simply mixing single-stranded oligonucleotides (ss-oligo), quaternized dextran (Q-dextran), and poly(l-lysine) (PLL) together. The resulting biphasic droplet, with PLL/ss-oligo phase being the internal subcompartments and Q-dextran/ss-oligo phase as the surrounding medium, is capable of sequestering and partitioning biomolecules into distinct regions. When the droplet is exposed to salt or Dextranase, the surrounding Q-dextran/ss-oligo phase will be gradually dissociated, resulting in the chaotic movement and fusion of internal subcompartments. Besides, the external electric field at a lower strength can drive the biphasic droplet to undergo a deviated circulation concomitant with the fusion of localized subcompartments, while a high-strength electric field can polarize the whole droplet, resulting in the release of daughter droplets in a controllable manner. Our study highlights that liquid-liquid phase separation of biopolymers is a powerful strategy to construct hierarchically structured protocells resembling the morphology and functions of living cells and provides a step toward a better understanding of the transition mechanism from nonliving to living matter under prebiotic conditions.

Mass Transport in Coacervate-Based Protocell Coated with Fatty Acid under Nonequilibrium Conditions.

Construction of protocell models from prebiotically plausible components to mimic the basic features or functions of living cells is still a challenge. In this work, we prepare a hybrid protocell model by coating sodium oleate on the coacervate droplet constituted by poly(l-lysine) and oligonucleotide and investigate the transport of different molecules under electric field. Results show that sodium oleate forms a layered viscoelastic membrane on the droplet surface, which is selectively permeable to small, polar molecules, such as oligolysine. As the droplet is stimulated at 10 V cm-1, the oleate membrane slips along the direction of electric field while maintaining its integrity. Most of the molecules are still excluded under such conditions. As repetitive cycles of vacuolization occur at 20 V cm-1, all molecules are internalized and sequestrated in the droplet through their specific pathways except enzyme, which anchors in the oleate membrane and is immune to electric field. Cascade enzymatic reactions are then carried out, and the product generated from the membrane exhibits a time-dependent concentration gradient across the droplet. Our work makes a step toward the nonequilibrium functionalization of synthetic protocells capable of biomimetic operations.

Genetic-based dissection of arsenic accumulation in maize using a genome-wide association analysis method.

Understanding the mechanism of arsenic (As) accumulation in plants is important in reducing As's toxicity to plants and its potential risks to human health. Here, we performed a genome-wide association study to dissect the genetic basis of the As contents of different maize tissues in Xixian, which was irrigated with As-rich surface water, and Changge using an association population consisting of 230 representative maize inbred lines. Phenotypic data revealed a wide normal distribution and high repeatability for the As contents in maize tissues. The As concentrations in maize tissues followed the same trend in the two locations: kernels < axes < stems < bracts < leaves. In total, 15, 16 and 15 non-redundant quantitative trait loci (QTLs) associated with As concentrations were identified (P ≤ 2.04 × 10-6 ) in five tissues from Xixian, Changge, and the combination of the locations, respectively, explaining 9.70%-24.65% of the phenotypic variation for each QTL, on average. Additionally, four QTLs [involving 15 single nucleotide polymorphisms (SNPs)] were detected in the single and the combined locations, indicating that these loci/SNPs might be stable across different environments. The candidate genes associated with these four loci were predicted. In addition, four non-redundant QTLs (6 SNPs), including a QTL that was detected in multiple locations according to the genome-wide association study, were found to co-localize with four previously reported QTL intervals. These results are valuable to understand the genetic architecture of As mechanism in maize and facilitate the genetic improvement of varieties without As toxicity.

External validation of different predictive scores for symptomatic intracranial hemorrhage after intravenous thrombolysis in Asian stroke patients.

OBJECTIVE: This study aimed to externally validate different predictive scores for symptomatic intracranial hemorrhage (SICH) after intravenous thrombolysis (IVT), with a particular focus on their predictive abilities in Asian stroke patients. METHODS: We retrospectively enrolled stroke patients who received a standard dose of alteplase within 4.5 hours from symptom onset at the First Affiliated Hospital of Dalian Medical University from July 2010 to August 2023. SICH was defined as the hemorrhagic transformation detected on the head CT scan completed within 48 h post-IVT, accompanied by a clinical deterioration of at least a 4-point increase in NIHSS score. Predictive abilities of the HAT, MSS, SEDAN, SPAN-100, and GRASPS scores were tested. Discrimination and calibration were performed using the area under the receiver operating characteristic curve (ROC-AUC), DeLong test, and Hosmer-Lemeshow (H-L) goodness-of-fit test. RESULTS: The study included 1007 stroke patients, of whom 31 (3.08 %) developed SICH. ROC-AUCs for predicting SICH were: 0.796 (95 %CI: 0.726-0.866) for the GRASPS score, 0.724 (95 %CI: 0.644-0.804) for the MSS score, 0.715 (95 %CI: 0.619-0.811) for the SEDAN score, 0.714 (95 %CI: 0.611-0.817) for the HAT score, and 0.605 (95 %CI: 0.491-0.720) for the SPAN-100 score (all P < 0.05). DeLong tests showed that the GRASPS score demonstrated significantly better discrimination than the MSS score (P = 0.010), the SEDAN score (P = 0.009), the HAT score (P = 0.049), and the SPAN-100 score (P = 0.000). H-L tests indicated good calibrations which were ranked HAT > SEDAN > MSS > SPAN-100 > GRASPS scores. CONCLUSION: The GRASPS score showed reasonable predictive ability for SICH, indicating its potential utility for Asian stroke patients receiving IVT.

Dual toeholds regulated CRISPR-Cas12a sensing platform for ApoE single nucleotide polymorphisms genotyping.

Single nucleotide polymorphisms (SNPs) are closely associated with many biological processes, including genetic disease, tumorigenesis, and drug metabolism. Accurate and efficient SNP determination has been proved pivotal in pharmacogenomics and diagnostics. Herein, a universal and high-fidelity genotyping platform is established based on the dual toeholds regulated Cas12a sensing methodology. Different from the conventional single stranded or double stranded activation mode, the dual toeholds regulated mode overcomes protospacer adjacent motif (PAM) limitation via cascade toehold mediated strand displacement reaction, which is highly universal and ultra-specific. To enhance the sensitivity for biological samples analysis, a modified isothermal recombinant polymerase amplification (RPA) strategy is developed via utilizing deoxythymidine substituted primer and uracil-DNA glycosylase (UDG) treatment, designated as RPA-UDG. The dsDNA products containing single stranded toehold domain generated in the RPA-UDG allow further incorporation with dual toeholds regulated Cas12a platform for high-fidelity human sample genotyping. We discriminate all the single-nucleotide polymorphisms of ApoE gene at rs429358 and rs7412 loci with human buccal swab samples with 100% accuracy. Furthermore, we engineer visual readout of genotyping results by exploiting commercial lateral flow strips, which opens new possibilities for field deployable implementation.