LncRNA AFAP1-AS1 mediates breast cancer cell proliferation and migration through the miR-21/PTEN axis.

LncRNAs play various effects, mostly by sponging with miRNAs. Based on public databases integrating bioinformatics analyses and further validation in breast cancer (BC) tissue and cell lines, the effect of lncRNA AFAP1-AS1 on breast cancer cell proliferation and migration was verified. It might work via the miR-21/PTEN axis. The expression of AFAP1-AS1, which was significantly upregulated in BC tissues and cell lines, was correlated with old age and lymph node metastasis of patients with BC. Knockdown of AFAP1-AS1 inhibited the proliferation and migration of BC cells in vitro and in vivo. And downregulated miR-21 expression and upregulated PTEN expression additionally. Mechanistically, the knockdown of lncRNA AFAP1-AS1 upregulated PTEN expression and consequently attenuated miR-21-mediated enhanced BC cell proliferation and migration. LncRNA AFAP1-AS1 is a potential prognostic biomarker for BC patients.

Chromosome-level genome assembly of Murraya paniculata sheds light on biosynthesis of floral volatiles.

BACKGROUND: Murraya paniculata (L.) Jack, commonly called orange jessamine in the family Rutaceae, is an important ornamental plant in tropical and subtropical regions which is famous for its strong fragrance. Although genome assemblies have been reported for many Rutaceae species, mainly in the genus Citrus, full genomic information has not been reported for M. paniculata, which is a prerequisite for in-depth genetic studies on Murraya and manipulation using genetic engineering techniques. Here, we report a high-quality chromosome-level genome assembly of M. paniculata and aim to provide insights on the molecular mechanisms of flower volatile biosynthesis. RESULTS: The genome assembly with a contig N50 of 18.25 Mb consists of 9 pseudomolecules and has a total length of 216.86 Mb. Phylogenetic analysis revealed that M. paniculata diverged from the common ancestor approximately 25 million years ago and has not undergone any species-specific whole genome duplication events. Genome structural annotation and comparative genomics analysis revealed that there are obvious differences in transposon contents among the genomes of M. paniculata and Citrus species, especially in the upstream regions of genes. Research on the flower volatiles of M. paniculata and C. maxima at three flowering stages revealed significant differences in volatile composition with the flowers of C. maxima lacking benzaldehyde and phenylacetaldehyde. Notably, there are transposons inserted in the upstream region of the phenylacetaldehyde synthase (PAAS) genes Cg1g029630 and Cg1g029640 in C. maxima, but not in the upstream region of three PAAS genes Me2G_2379, Me2G_2381, and Me2G_2382 in M. paniculata. Our results indicated that compared to the low expression levels of PAAS genes in C. maxima, the higher expression levels of the three PAAS genes in M. paniculata are the main factor affecting the phenylacetaldehyde biosynthesis and causing the content difference of phenylacetaldehyde. The phenylacetaldehyde synthetic activities of the enzymes encoded by M. paniculata PAAS genes were validated by in vitro analyses. CONCLUSIONS: Our study provides useful genomic resources of M. paniculata for further research on Rutaceae plants, identifies new PAAS genes, and provides insights into how transposons contribute to variations in flower volatiles among Murraya and Citrus plants.

ICE1 interacts with IDD14 to transcriptionally activate QQS to increase pollen germination and viability.

In flowering plants, sexual reproductive success depends on the production of viable pollen grains. However, the mechanisms by which QUA QUINE STARCH (QQS) regulates pollen development and how transcriptional activators facilitate the transcription of QQS in this process remain poorly understood. Here, we demonstrate that INDUCER OF CBF EXPRESSION 1 (ICE1), a basic helix-loop-helix (bHLH) transcription factor, acts as a key transcriptional activator and positively regulates QQS expression to increase pollen germination and viability in Arabidopsis thaliana by interacting with INDETERMINATE DOMAIN14 (IDD14). In our genetic and biochemical experiments, overexpression of ICE1 greatly promoted both the activation of QQS and high pollen viability mediated by QQS. IDD14 additively enhanced ICE1 function by promoting the binding of ICE1 to the QQS promoter. In addition, mutation of ICE1 significantly repressed QQS expression; the impaired function of QQS and the abnormal anther dehiscence jointly affected pollen development of the ice1-2 mutant. Our results also showed that the enhancement of pollen activity by ICE1 depends on QQS. Furthermore, QQS interacted with CUT1, the key enzyme for long-chain lipid biosynthesis. This interaction both promoted CUT1 activity and regulated pollen lipid metabolism, ultimately determining pollen hydration and fertility. Our results not only provide new insights into the key function of QQS in promoting pollen development by regulating pollen lipid metabolism, but also elucidate the mechanism that facilitates the transcription of QQS in this vital developmental process.

B13, a well-tolerated inhibitor of hedgehog pathway, exhibited potent anti-tumor effects against colorectal carcinoma in vitro and in vivo.

Abnormal activation of Hedgehog (Hh) signaling pathway mediates the genesis and progression of various tumors [1]. Currently, three drugs targeting the Hh signaling component Smoothened (Smo) have been marketed for the clinical treatment of basal cell tumors or acute myeloid leukemia. However, drug resistance is a common problem in those drugs, so the study of Smo inhibitors that can overcome drug resistance has important guiding significance for clinical adjuvant drugs. MTT assay, clone formation assay and EdU assay were used to detect the proliferation inhibitory activity of the drugs on tumor cells. The effect of B13 on cell cycle and apoptosis were detected by flow cytometry. An acute toxicity test was used to detect the toxicity of B13 in vivo, and xenograft tumor model was used to detect the efficacy of B13 in vivo. The binding of B13 to Smo was studied by BODIPY-cyclopamine competitive binding assay and molecular docking. The effect of B13 on the expression and localization of downstream target gene Gli1/2 of Smo was investigated by Western Blot and immunofluorescence assay. SmoD473H mutant cell line was constructed to study the effect of B13 against drug resistance. (1) B13 had the strongest inhibitory activity against colorectal cancer cells. (2) B13 can effectively inhibit the clone formation and EdU positive rate of colon cancer cells. (3) B13 can block the cell cycle in the G2/M phase and cell apoptosis. (4) B13 has low toxicity in vivo, and its efficacy in vivo is better than that of the Vismodegib. (5) Molecular docking and BODIPY-cyclopamine experiments showed that B13 could bind to Smo protein. (6) B13 can inhibit the protein expression of Gli1, the downstream of Smo, and inhibit its entry into the nucleus. (7) B13 could inhibit the expression of Gli1 in the HEK293 cells with SmoD473H, and the molecular docking results showed that B13 could bind SmoD473H protein. B13 with the best anti-tumor activity was screened out by MTT assay. In vitro, pharmacodynamics experiments showed that B13 could effectively inhibit the proliferation and metastasis of colorectal cancer cells, induce cell cycle arrest, and induce cell apoptosis. In vivo pharmacodynamics experiments showed that B13 was superior to Vismodegib in antitumor activity and had low toxicity in vivo. Mechanism studies have shown that B13 can bind Smo protein, inhibit the expression of downstream Gli1 and its entry into the nucleus. Notably, B13 overcomes resistance caused by SmoD473H mutations.

Real-world effectiveness of benralizumab: Results from the ZEPHYR 1 Study.

BACKGROUND: Real-world evidence characterizing the clinical outcomes and economic impact on patients with severe eosinophilic asthma treated with benralizumab is limited. OBJECTIVE: To characterize patients with severe asthma treated with benralizumab and assess its clinical and economic impact in the United States. METHODS: A pre-post benralizumab comparison was performed using a large US insurance claims database between November 2016 and November 2019. The primary cohort included patients with asthma aged 12 years or more with 2 or more records of benralizumab. Secondary cohorts included persistent users (6 or more records of benralizumab), patients switching to benralizumab from mepolizumab or omalizumab, and stratified by Medicaid vs non-Medicaid. Exacerbations, concomitant medications, and exacerbation-related health care resource utilization (HCRU) and costs were compared in the 12-month periods pre- and post-benralizumab initiation (index). RESULTS: Of the 204 patients in the primary cohort, mean age at index was 45.3 years and 68.6% were of female sex. The patients experienced a significant 55% reduction in rates of exacerbations post-benralizumab initiation (3.25 pre-index vs 1.47 post-index per person-year; P < .001), and 41% of the patients had no exacerbations post-benralizumab initiation. The proportion of oral corticosteroid-dependent patients decreased from 82% to 50% (P < .001). Reductions in HCRU were 42%, 46%, and 57% for asthma exacerbation-related inpatient hospitalizations, emergency department, and outpatient visits, respectively (all P < .001). Exacerbation-related costs decreased by $6439 ($13,559 vs $7120; P < .001). Similar results for all outcomes were observed for the persistent cohort, switch cohorts, and Medicaid vs non-Medicaid cohorts. CONCLUSION: Patients with severe asthma treated with benralizumab experienced clinical and economic benefits in the real world, as demonstrated by the reduction in exacerbations and HCRU.

Cold Response Transcriptome Analysis of the Alternative Splicing Events Induced by the Cold Stress in D. catenatum.

Dendrobium catenatum Lindl is a valuable medicinal herb and gardening plant due to its ornamental value and special medical value. Low temperature is a major bottleneck restricting D. catenatum expansion towards the north, which influences the quality and yield of D. catenatum. In this study, we analysed the cold response of D. catenatum by RNA-Seq. A total of 4302 differentially expressed genes were detected under cold stress, which were mainly linked to protein kinase activity, membrane transport and the glycan biosynthesis and metabolism pathway. We also identified 4005 differential alternative events in 2319 genes significantly regulated by cold stress. Exon skipping and intron retention were the most common alternative splicing isoforms. Numerous genes were identified that differentially modulated under cold stress, including cold-induced transcription factors and splicing factors mediated by AS (alternative splicing). GO enrichment analysis found that differentially alternatively spliced genes without differential expression levels were related to RNA/mRNA processing and spliceosomes. DAS (differentially alternative splicing) genes with different expression levels were mainly enriched in protein kinase activity, plasma membrane and cellular response to stimulus. We further identified and cloned DcCBP20 in D. catenatum; we found that DcCBP20 promotes the generation of alternative splicing variants in cold-induced genes under cold stress via genetic experiments and RT-PCR. Taken together, our results identify the main cold-response pathways and alternative splicing events in D. catenatum responding to cold treatment and that DcCBP20 of D. catenatum get involved in regulating the AS and gene expression of cold-induced genes during this process. Our study will contribute to understanding the role of AS genes in regulating the cold stress response in D. catenatum.

Enhanced antitumor efficacy by combining afatinib with MDV3100 in castration-resistant prostate cancer.

Background: Patients with prostate cancer often develop resistance to androgen deprivation therapy, a condition called castration-resistant prostate cancer (CRPC). Enzalutamide (MDV3100) can prolong the survival of patients with CRPC after chemotherapy, but ∼50% of patients eventually relapse and develop resistance to MDV3100. Thus, it is necessary to explore new treatment methods to improve the therapeutic effect of MDV3100. Tyrosine kinases play an essential role in the pathogenesis of CRPC. Methods: MTT assay was used to detect the inhibitory effects of MDV3100 and tyrosine kinase inhibitor on prostate cancer cells. CompuSyn version 1.0 was used to calculate the combination index (CI) values using the Chou-Talalay method. Clone formation and EdU assay were used to detect the effect of afatinib combined with MDV3100 on the proliferation of 22Rv1 cells. RT-qPCR and Western blot were used to explore the mechanism of drug combination. The aim of the present study was to determine the effects of several tyrosine kinase inhibitors (TKIs) when used in combination with MDV3100 in vitro. Results: The results demonstrated that TKIs combined with MDV3100 exerted a synergistic effect on a variety of PCa cells. Afatinib combined with MDV3100 could suppress the proliferation and migration of 22RV1 cells, as well as cause their cell cycle arrest and apoptosis. Mechanistically, afatinib effectively reduced the protein expression levels of HER2 and HER3 and inhibited EGFR phosphorylation, thereby enhancing the effect of MDV3100 and suppressing CRPC. Conclusions: These findings suggested that afatinib treatment improved the effect of MDV3100 on 22RV1 cells, highlighting this drug as a potential therapeutic strategy for patients with CRPC.

Ergosterol-targeting fusion antifungal peptide significantly increases the Verticillium wilt resistance of cotton.

Increasing the targeting ability of antifungal proteins towards specific components of fungal cells has the potential to improve their antifungal activity and reduce harmful effects to nontarget cells. To obtain effective disease resistance genes against cotton Verticillium wilt, we constructed several fusion genes, in which binding domains targeting chitin, sphingolipid or ergosterol in the fungal cell wall or cell membrane were individually fused to the antifungal peptide BbAFP1 from entomopathogenic fungus Beauveria bassiana. Transient expression of fusion genes in cotton cotyledons indicated that the BbAFP1::ErBD fusion peptide with an ergosterol binding domain exhibited better disease resistance against V. dahliae than wild-type BbAFP1 and other fusion genes. BbAFP1::ErBD and BbAFP1 transgenic cotton were obtained and verified by Southern and Western blotting. Compared with BbAFP1-expressing cotton, BbAFP1::ErBD-expressing cotton showed higher disease resistance against V. dahliae, with smaller lesion areas (0.07 cm2 vs. 0.16 cm2 ) on the leaves and a lower disease index (23.9 vs. 34.5). Overexpression of BbAFP1::ErBD by transgenic tobacco also showed enhanced disease resistance against V. dahliae compared with that of the wild-type gene. These results indicated that construction of fusion antifungal peptides that target fungal cells is a powerful strategy to obtain new anti-disease genes, and the obtained fusion gene BbAFP1::ErBD has the potential to defend against plant fungal diseases.

Genome-Wide Analysis of the TCP Transcription Factor Genes in Dendrobium catenatum Lindl.

Teosinte branched1/cycloidea/proliferating cell factor (TCP) gene family members are plant-specific transcription factors that regulate plant growth and development by controlling cell proliferation and differentiation. However, there are no reported studies on the TCP gene family in Dendrobium catenatum Lindl. Here, a genome-wide analysis of TCP genes was performed in D. catenatum, and 25 TCP genes were identified. A phylogenetic analysis classified the family into two clades: Class I and Class II. Genes in the same clade share similar conserved motifs. The GFP signals of the DcaTCP-GFPs were detected in the nuclei of tobacco leaf epidermal cells. The activity of DcaTCP4, which contains the miR319a-binding sequence, was reduced when combined with miR319a. A transient activity assay revealed antagonistic functions of Class I and Class II of the TCP proteins in controlling leaf development through the jasmonate-signaling pathway. After different phytohormone treatments, the DcaTCP genes showed varied expression patterns. In particular, DcaTCP4 and DcaTCP9 showed opposite trends after 3 h treatment with jasmonate. This comprehensive analysis provides a foundation for further studies on the roles of TCP genes in D. catenatum.

Functional Characterization of the Stipa purpurea P5CS Gene under Drought Stress Conditions.

Free proline has multiple functions in plant cells, such as regulating osmotic potential and protecting both proteins and cell membranes. The expression of Δ1-Pyrroline-5-carboxylate synthase (P5CS), a key enzyme in the proline biosynthetic pathway, increases under drought, salt and cold stress conditions, causing plant cells to accumulate large amounts of proline. In this study, we cloned and identified the P5CS gene from Stipa purpurea, which has a full-length of 2196 bp and encodes 731 amino acids. A subcellular localization analysis indicated that SpP5CS localized to the cytoplasm. The ectopic overexpression of SpP5CS in Arabidopsis thaliana resulted in higher proline contents, longer roots, higher survival rates and less membrane damage under drought stress conditions compared with wild-type controls. SpP5CS-overexpressing A. thaliana was more resistant to drought stress than the wild type, whereas the deletion mutant sp5cs was less resistant to drought stress. Thus, SpP5CS may be a potential candidate target gene for increasing plant resistance to drought stress.

Genome-Wide Identification of OSC Gene Family and Potential Function in the Synthesis of Ursane- and Oleanane-Type Triterpene in Momordica charantia.

The triterpenes in bitter gourd (Momordica charantia) show a variety of medicinal activities. Oxidosqualene cyclase (OSC) plays an indispensable role in the formation of triterpene skeletons during triterpene biosynthesis. In this study, we identified nine genes encoding OSCs from bitter gourd (McOSC1-9). Analyses of their expression patterns in different tissues suggested that characteristic triterpenoids may be biosynthesized in different tissues and then transported. We constructed a hairy root system in which McOSC7 overexpression led to an increased accumulation of camaldulenic acid, enoxolone, and quinovic acid. Thus, the overexpression of McOSC7 increased the active components content in bitter gourd. Our data provide an important foundation for understanding the roles of McOSCs in triterpenoid synthesis.

Distributed Deep Fusion Predictor for a Multi-Sensor System Based on Causality Entropy.

Trend prediction based on sensor data in a multi-sensor system is an important topic. As the number of sensors increases, we can measure and store more and more data. However, the increase in data has not effectively improved prediction performance. This paper focuses on this problem and presents a distributed predictor that can overcome unrelated data and sensor noise: First, we define the causality entropy to calculate the measurement's causality. Then, the series causality coefficient (SCC) is proposed to select the high causal measurement as the input data. To overcome the traditional deep learning network's over-fitting to the sensor noise, the Bayesian method is used to obtain the weight distribution characteristics of the sub-predictor network. A multi-layer perceptron (MLP) is constructed as the fusion layer to fuse the results from different sub-predictors. The experiments were implemented to verify the effectiveness of the proposed method by meteorological data from Beijing. The results show that the proposed predictor can effectively model the multi-sensor system's big measurement data to improve prediction performance.

Identification of lncRNA NEAT1/miR-21/RRM2 axis as a novel biomarker in breast cancer.

As a major disease that threatens the health of women worldwide, breast cancer (BC) lacks effective molecular markers in the clinic at the same time. We aim at finding a new biomarker of BC. In our study, through the Gene Expression Omnibus database chip, a total of 1393 pairs of microRNA-messenger RNA (miRNA-mRNA) networks and 35754 pairs of long noncoding RNA-miRNA networks were obtained. We found out that NEAT1/miR-21/RRM2 axis may play a role in BC diagnosis and prognosis. The real-time quantitative reverse transcription-polymerase chain reaction test was used to analyze the mRNA level of NEAT1, miR-21, and RRM2. Western blot was used to detect the protein level of RRM2. Through the 5-ethynyl-2'-deoxyuridine assay, the proliferation of MDA-MB-231 cells was detected. Through wound healing and transwell assay, the migration of MDA-MB-231 cells was detected. Altogether, our data indicated that NEAT1, miR-21, and RRM2 were upregulated in several BC cell lines. Overexpressed of miR-21 in MDA-MB-231 cells promote proliferation and migration. Besides, our results demonstrated that overexpressed of miR-21 upregulated the level of RRM2. Accordingly, miR-21/RRM2 might be a new diagnosis and treatment target of BC.

Comparative transcriptome analysis reveals ecological adaption of cold tolerance in northward invasion of Alternanthera philoxeroides.

BACKGROUND: Alternanthera philoxeroides (alligator weed) is a highly invasive alien plant that has continuously and successfully expanded from the tropical to the temperate regions of China via asexual reproduction. During this process, the continuous decrease in temperature has been a key limiting environmental factor. RESULTS: In this study, we provide a comprehensive analysis of the cold tolerance of alligator weed via transcriptomics. The transcriptomic differences between the southernmost population and the northernmost population of China were compared at different time points of cold treatments. GO enrichment and KEGG pathway analyses showed that the alligator weed transcriptional response to cold stress is associated with genes encoding protein kinases, transcription factors, plant-pathogen interactions, plant hormone signal transduction and metabolic processes. Although members of the same gene family were often expressed in both populations, the levels of gene expression between them varied. Further ChIP experiments indicated that histone epigenetic modification changes at the candidate transcription factor gene loci are accompanied by differences in gene expression in response to cold, without variation in the coding sequences of these genes in these two populations. These results suggest that histone changes may contribute to the cold-responsive gene expression divergence between these two populations to provide the most beneficial response to chilling stimuli. CONCLUSION: We demonstrated that the major alterations in gene expression levels belonging to the main cold-resistance response processes may be responsible for the divergence in the cold resistance of these two populations. During this process, histone modifications in cold-responsive genes have the potential to drive the major alterations in cold adaption necessary for the northward expansion of alligator weed.

BrrICE1.1 is associated with putrescine synthesis through regulation of the arginine decarboxylase gene in freezing tolerance of turnip (Brassica rapa var. rapa).

BACKGROUND: In the agricultural areas of Qinghai-Tibet Plateau, temperature varies widely from day to night during the growing season, which makes the extreme temperature become one of the limiting factors of crop yield. Turnip (Brassica rapa var. rapa) is a traditional crop of Tibet grown in the Tibet Plateau, but its molecular and metabolic mechanisms of freezing tolerance are unclear. RESULTS: Here, based on the changes in transcriptional and metabolic levels of Tibetan turnip under freezing treatment, the expression of the arginine decarboxylase gene BrrADC2.2 exhibited an accumulative pattern in accordance with putrescine content. Moreover, we demonstrated that BrrICE1.1 (Inducer of CBF Expression 1) could directly bind to the BrrADC2.2 promoter, activating BrrADC2.2 to promote the accumulation of putrescine, which was verified by RNAi and overexpression analyses for both BrrADC2.2 and BrrICE1.1 using transgenic hair root. The function of putrescine in turnip was further analyzed by exogenous application putrescine and its inhibitor DL-α-(Difluoromethyl) arginine (DFMA) under freezing tolerance. In addition, the BrrICE1.1 was found to be involved in the ICE1-CBF pathway to increase the freezing stress of turnip. CONCLUSIONS: BrrICE1.1 could bind the promoter of BrrADC2.2 or CBFs to participate in freezing tolerance of turnip by transcriptomics and targeted metabolomics analyses. This study revealed the regulatory network of the freezing tolerance process in turnip and increased our understanding of the plateau crops response to extreme environments in Tibet.

Uncovering the role of a positive selection site of wax ester synthase/diacylglycerol acyltransferase in two closely related Stipa species in wax ester synthesis under drought stress.

Natural selection drives local adaptations of species to biotic or abiotic environmental stresses. As a result, adaptive phenotypic divergence can evolve among related species living in different habitats. However, the genetic foundation of this divergence process remains largely unknown. Two closely related alpine grass species, Stipa capillacea and Stipa purpurea, are distributed in different rainfall regions of northern Tibet. Here, we analyzed the drought tolerance of these two closely related Stipa species, and found that S. purpurea was more resistance to drought stress than S. capillacea. To further understand the genetic diversity behind their adaptation to drought environments, a comprehensive gene repertoire was generated using PacBio isoform and Illumina RNA sequencing technologies. Bioinformatics analyses revealed that differential transcripts were mainly enriched in the wax synthetic pathway, and a threonine residue at position 239 of WSD1 was identified as having undergone positive selection in S. purpurea. Using heterologous expression in the Saccharomyces cerevisiae mutant H1246, site-directed mutagenesis studies demonstrated that a positive selection site results in changes to the wax esters profile. This difference may play an important role in S. purpurea in response to drought conditions, indicating that S. purpurea has evolved specific strategies involving its wax biosynthetic pathway as part of its long-term adaptation to the Qinghai-Tibet Plateau.

Discovery of [1,2,4]triazolo[4,3-a]pyridines as potent Smoothened inhibitors targeting the Hedgehog pathway with improved antitumor activity in vivo.

Triple-negative breast cancer (TNBC), a subset of breast cancers, have poorer survival than other breast cancer types. Recent studies have demonstrated that the abnormal Hedgehog (Hh) pathway is activated in TNBC and that these treatment-resistant cancers are sensitive to inhibition of the Hh pathway. Smoothened (Smo) protein is a vital constituent in Hh signaling and an attractive drug target. Vismodegib (VIS) is one of the most widely studied Smo inhibitors. But the clinical application of Smo inhibitors is limited to adult patients with BCC and AML, with many side effects. Therefore, it's necessary to develop novel Smo inhibitor with better profiles. Twenty [1,2,4]triazolo[4,3-a]pyridines were designed, synthesized and screened as Smo inhibitors. Four of these novel compounds showed directly bound to Smo protein with stronger binding affinity than VIS. The new compounds showed broad anti-proliferative activity against cancer cell lines in vitro, especially triple-negative breast cancer cells. Mechanistic studies demonstrated that TPB15 markedly induced cell cycle arrest and apoptosis in MDA-MB-468 cells. TPB15 blocked Smo translocation into the cilia and reduced Smo protein and mRNA expression. Furthermore, the expression of the downstream regulatory factor glioma-associated oncogene 1 (Gli1) was significantly inhibited. Finally, TPB15 demonstrated greater anti-tumor activity in our animal models than VIS with lower toxicity. Hence, these results support further optimization of this novel scaffold to develop improved Smo antagonists.

Carboxyl Fe3O4 magnetic nanoparticle-based SPE and HPLC method for the determination of six tetracyclines in water.

A novel carboxyl Fe3O4 magnetic nanoparticle-based solid-phase extraction combined with high-performance liquid chromatography was developed for the analysis of oxytetracycline, tetracycline, demeclocycline, metacycline, chlortetracycline, and doxycycline in water samples. Driven by the electrostatic interaction and the strong chelation between tetracyclines and iron ions, tetracyclines in samples were adsorbed onto the adsorbents. The adsorbed analytes were subsequently eluted with oxalic acid and separated with a C18 column under gradient condition with a mobile phase consisting of methanol, acetonitrile, and oxalic acid at a flow rate of 0.5 mL/min. The detection was performed at variable ultraviolet wavelengths. Under optimized conditions, the developed method gave an enrichment factor of 33.3, linearity ranges of 5.00-1000 µg/L, detection limits of (2.86-5.19) × 10-2 µg/L, quantification limits of (9.54-17.3) × 10-2 µg/L, recoveries of 76.2-98.0%, and intra- and inter-day RSDs of 0.132-15.5% and 2.28-14.5% for these tetracyclines. The established method was successfully applied for the determination of these six tetracyclines in tap water, river water, pond water, and lake water samples. Graphical abstract ᅟ.

Molecular cloning and characterization of the glutathione reductase gene from Stipa purpurea.

Reactive oxygen species (ROS) are a key factor in abiotic stresses; excess ROS is harmful to plants. Glutathione reductase (GR) plays an important role in scavenging ROS in plants. Here, a GR gene, named SpGR, was cloned from Stipa purpurea and characterized. The full-length open reading frame was 1497 bp, encoding 498 amino acids. Subcellular localization analysis indicated that SpGR was localized to both the plasma membrane and nucleus. The expression of SpGR was induced by cold, salt, and drought stresses. Functional analysis indicated that ectopic expression of SpGR in Arabidopsis thaliana resulted in greater tolerance to salt stress than that of wild-type plants, but no difference under cold or drought treatments. The results of GR activity and GSSG and GSH content analyses suggested that, under salt stress, transgenic plants produced more GR to reduce GSSG to GSH for scavenging ROS than wild-type plants. Therefore, SpGR may be a candidate gene for plants to resist abiotic stress.