Frontiers in Microbiology
·
2019
·
IF 4.259
He Shengnan,An Tai,A Runa,Liu Shouan
The tea plant [Camellia sinensis (L.) O. Kuntze] is one of the most important leaf crops, and it is widely used for the production of non-alcoholic beverages worldwide. Tea also has a long history of medicinal use. Colletotrichum camelliae Massee is one of the dominant fungal pathogens that infects tea leaves and causes severe tea anthracnose disease. To analyze the molecular biology of C. camelliae, the quantification of pathogen gene expression by the RT-qPCR method is necessary. Reliable RT-qPCR results require the use of stable reference genes for data normalization. However, suitable reference genes have not been reported in C. camelliae thus far. In this study, 12 candidate genes (i.e., CcSPAC6B12.04c, CcWDR83, Cchp11, Ccnew1, CcHplo, CcRNF5, CcHpcob, CcfaeB-2, CcYER010C, CcRNM1, CcUP18, and CcACT) were isolated from C. camelliae and assessed as potential reference genes. The expression stability of these genes in C. camelliae during spore germination and mycelial growth and interaction with host plants was first evaluated using several statistical algorithms, such as geNorm, NormFinder, and Bestkeeper. A web-based analysis program, Refinder, was then used to find the most suitable reference genes. Our results indicated that Cenew1, CcHplo, and CcSPAC6B12.04c were the most stable reference genes in C. camelliae under all conditions. Our work provided the most suitable reference genes for future studies performed to quantify the target gene expression levels of C. camelliae.
研究领域:
植物病理学
·
分子生物学
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DOI
PubMed
高影响因子
Analytical Methods
·
2024
·Vol. 16
·期 8
·1281-1287
·
IF 2.7
Keyu Li,Peng Xiao,Ningning Yuan,Shujuan Yan,Pei Zhao,Guowei Zuo
MiRNAs are critical regulators of target gene expression in many biological processes and are considered promising biomarkers for diseases. In this study, we developed a simple, specific, and sensitive miRNA detection method based on proximity ligation reaction, which is easy to operate. The method uses a pair of target-specific DNA probes immobilized on the same gold nanoparticles (AuNPs), which hybridize to the target miRNA. Hybridization brings the probes close together, allowing the formation of a continuous DNA sequence that can be amplified by Quantitative Real-time PCR (qPCR). This method eliminates the need for complex reverse transcription design and achieves high specificity for discriminating single base mismatches between miRNAs through a simple procedure. This method can sensitively measure three different miRNAs with a detection limit of 20 aM, providing high versatility and sensitivity, even distinguishing single-base variations among members of the miR-200 family with high selectivity. Due to its high selectivity and sensitivity, this method has important implications for the investigation of miRNA biological functions and related biomedical research.
研究领域:
分子生物学
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纳米生物技术
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医学检测
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产品关联
DOI
PubMed
高影响因子
JOURNAL OF NANOPARTICLE RESEARCH
·
2014
·Vol. 16
·期 11
·1-13
·
IF 2.278
Liu Wenjuan,Yao Jun,Cai Minmin,Chai Hankuai,Zhang Chi,Sun Jingjing,Chandankere Radhika,Masakorala Kanaji
A new nanopesticide, carboxymethyl-β-cyclodextrin-Fe 3 O 4 magnetic nanoparticles-Diuron (CM-β-CD-MNPs-Diuron), was synthesized from an inclusion complex of CM-β-CD-MNPs as host and diuron as guest molecules. The transmission electron microscopy revealed it had an average diameter of 25 nm which is more or less the same as that of MNPs (average diameter ~23 nm). The CM-β-CD grafting was confirmed by infrared spectroscopy, and the amount of CM-β-CD grafted on the surface of MNPs was determined to be 144.1 mg/g by thermogravimetry. The feasibility of using CM-β-CD-MNPs as a nanocarrier for loading diuron was verified by investigating the formation of inclusion complex. The complexation of CM-β-CD-MNPs with diuron followed the Langmuir adsorption isotherm. In this work, the potential toxic effect of CM-β-CD-MNPs-Diuron on soil microbial was evaluated by microcalorimetry, urease enzyme and real-time quantitative PCR (qPCR). The thermokinetic parameters were observed to decrease with increase in the loading of CM-β-CD-MNPs-Diuron in soil. The urease activity data showed that there was a significant effect ( p < 0.05) of CM-β-CD-MNPs-Diuron on the enzyme activity. The microcalorimetric analysis was in agreement with qPCR, confirming the toxic effect of this nanopesticide on microorganism in soil.
研究领域:
环境科学
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微生物学
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纳米技术
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产品关联
DOI
高影响因子
BMC PLANT BIOLOGY
·
2019
·Vol. 19
·期 1
·1-16
·
IF 3.67
Cao Yunpeng,Meng Dandan,Han Yahui,Chen Tianzhe,Jiao Chunyan,Chen Yu,Jin Qing,Cai Yongping
Background Studies have demonstrated that BBX (B-BOX) genes play crucial roles in regulatory networks controlling plant growth, developmental processes and stress response. Nevertheless, comprehensive study of BBX genes in orchids (Orchidaceae) is not well studied. The newly released genome sequences of Dendrobium officinale and Phalaenopsis equestris have allowed a systematic analysis of these important BBX genes in orchids. Results Here we identified 19 ( DoBBX01 – 19 ) and 16 ( PeBBX01–16 ) BBX genes from D. officinale and P. equestris , respectively, and clustered into five clades (I-V) according to phylogenetic analysis. Thirteen orthologous, two DoBBXs paralogous and two PeBBXs paralogous gene pairs were validated. This gene family mainly underwent purifying selection, but five domains experienced positive selection during evolution. Noteworthy, the expression patterns of root, root_tips, stem, leaf, speal, column, lip, and flower_buds revealed that they might contribution to the formation of these tissues. According to the cis-regulatory elements analysis of BBX genes, qRT-PCR experiments were carried out using D. officinale PLBs (protocorm-like bodies) and displayed that these BBX genes were differentially regulated under AgNO 3 , MeJA (Methyl Jasmonate), ABA (abscisic acid) and SA (salicylic acid) treatments. Conclusions Our analysis exposed that DoBBX genes play significant roles in plant growth and development, and response to different environmental stress conditions of D. officinale , which provide aid in the selection of appropriate candidate genes for further functional characterization of BBX genes in plants.
研究领域:
植物生物学
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基因组学
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分子遗传学
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生物信息学
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DOI
PubMed
高影响因子
Bioengineered
·
2021
·
IF 3.269
Jianan Li,Haoran Bi
Cypermethrin (CMN) is a widely used artificial synthetic pesticide that causes neurotoxicity in the hippocampus. However, the underlying toxicological targets and mechanisms remain unclear. In this study, network pharmacology analysis and in vitro models were integrated to investigate the effect and mechanism of CMN-induced hippocampal neurotoxicity. A total of 88 targets of CMN-induced hippocampal neurotoxicity were predicted. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes enrichment (KEGG) analyses suggested that these targets were related to multiple GO terms and signaling pathways. To further investigate underlying mechanism, the top 10 hub targets (Akt1, Tnf, Ptgs2, Casp3, Igf1, Sirt1, Jun, Cat, Il10, and Bcl2l1) were screened. Furthermore, cell viability and lactate dehydrogenase (LDH) assays demonstrated that CMN was toxic to HT22 cells in a time- and dose-dependent manner. Terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining revealed that treatment with CMN increased the proportion of apoptotic cells. In addition, the real-time quantitative polymerase chain reaction (RT-qPCR) results indicated that CMN altered the mRNA expression levels of most of the hub targets, with the exceptions of Igf1 and Jun. The results demonstrated that multiple targets and signaling pathways were involved in CMN-induced hippocampal neurotoxicity. These findings provided reference values for subsequent studies of the toxicological mechanism of CMN.
研究领域:
毒理学
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神经科学
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药理学
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DOI
PubMed
高影响因子
JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES
·
2024
·
IF 2.8
Yan-Long Zhang,Yu-Jun Shi,Bao-Jian Duan,Xian-Hua Wang
For separation of deoxyribonucleic acid (DNA), positively charged amino-modified magnetic nanoparticles (MN) can effectively adsorb negatively charged DNA through electrostatic interaction. However, the reported preparation of amino-modified MN is usually tedious and time-consuming. Therefore, a simple synthesis method of amino-modified MN is necessary for DNA extraction. Herein, a novel polyethyleneimine-coated MN (PMN) was fabricated by one-pot hydrothermal synthesis for high-efficient DNA extraction. The fabricated PMN showed numerous exposed amino groups, which not only could effectively capture DNA through electrostatic interaction, but also limited the aggregation of PMN during application. Under optimized adsorption conditions, the maximum adsorption capacity of PMN for DNA could reach 192.4 μg m g−1. Shigella flexneri ( S . flexneri ) has the highest mortality rate among Shigella species and has been selected as target model pathogenic bacteria. Based on the optimized extraction conditions, PMN-based magnetic solid-phase microextraction (MSPE) and quantitative real-time PCR (qPCR) were integrated for detection of S . flexneri . The limit of detection of the proposed strategy was 2.4 × 10 2 CFU mL −1 and obviously lower than the commercial kit. To prove the practicability, the PMN-based MSPE combined qPCR strategy was successfully used in the determination of S . flexneri in spiked real sample, and the recovery values were in the range of 95.1 % to 102.1 % for apple juice, 60.4 % to 85.7 % for pickled vegetable, 97.8 % to 99.5 % for pig liver and 95.1 % to 102.1 % for pig colon, respectively. Therefore, we believe that the resultant PMN have great potential to become a universal magnetic adsorbent for high-efficient DNA extraction from complex biological samples.
研究领域:
生物化学
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分子生物学
·
医学检测
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产品关联
DOI
PubMed
高影响因子
INTERNATIONAL JOURNAL OF MOLECULAR SCIENCES
·
2019
·Vol. 20
·期 11
·2694
·
IF 4.183
Wenqi Li,Yubing Yong,Yue Zhang,Yingmin Lyu
Background: The LA hybrid lily ‘Aladdin’ has both excellent traits of Longiflorum hybrids and Asiatic hybrids—such as big and vivid flower, strong stem, high self-propagation coefficient, and shorter low temperature time required to release bulb dormancy in contrast to Oriental hybrids. A genome-wide transcriptional analysis using transcriptome RNA-Seq was performed in order to explore whether there is a gibberellin floral induction pathway in the LA hybrid lily. Subsequently, gene co-expression network analysis was used to analyze the possible interactions of key candidate genes screened from transcriptome data. At the same time, a series of physiological, biochemical, and cultivation tests were carried out. Results: The content of five endogenous hormones changed sharply in the shoot apex during the treatment of 200 mg/L exogenous gibberellin and the ratio of ABA/GA3dropped and stayed at a lower level after 4 hours’ treatment from the higher levels initially, reaching a dynamic balance. In addition, the metabolism of carbohydrates in the bulbs increase during exogenous gibberellin treatment. A total of 124,041 unigenes were obtained by RNA-seq. With the transcriptome analysis, 48,927 unigenes and 48,725 unigenes respectively aligned to the NR database and the Uniprot database. 114,138 unigenes, 25,369 unigenes, and 19,704 unigenes respectively aligned to the COG, GO, and KEGG databases. 2148 differentially expression genes (DEGs) were selected with the indicators RPKM ≥ 0, FDR ≤ 0.05 and |log2(ratio)| ≥ 2. The number of the upregulated unigenes was significantly more than the number of the downregulated unigenes. Some MADS-box genes related to flowering transformation—such asAGL20,SOC1, andCO—were found to be upregulated. A large number of gibberellin biosynthesis related genes such asGA2ox,GA3ox,GA20ox,Cytochrome P450,CYP81, and gibberellin signal transduction genes such asDELLA,GASA, andGID1were significantly differentially expressed. The plant hormones related genes such asNCED3and sugar metabolism related genes such asα-amylase,sucrose synthase hexokinase, and so on were also found expressing differentially. In addition, stress resistance related genes such asLEA1,LEA2,LEA4,serine/threonine protein kinase,LRR receptor-like serine/threonine protein kinase,P34 kinase,histidine kinase 3and epigenetic related genes in DNA methylation, histone methylation, acetylation, ubiquitination of ribose were also found. Particularly, a large number of transcription factors responsive to the exogenous gibberellin signal includingWRKY40, WRKY33, WRKY27, WRKY21, WRKY7, MYB, AP2/EREBP, bHLH, NAC1, NAC2, andNAC11were found to be specially expressing. 30 gene sequences were selected from a large number of differentially expressed candidate genes for qRT-PCR expression verification (0, 2, 4, 8, and 16 h) and compared with the transcriptome expression levels. Conclusions: 200mg/L exogenous GA3can successfully break the bulb’s dormancy of the LA hybrid lily and significantly accelerated the flowering process, indicating that gibberellin floral induction pathway is present in the LA lily ‘Aladdin’. With the GCNs analysis, two second messenger G protein-coupled receptor related genes that respond to gibberellin signals in the cell were discovered. The downstream transport proteins such asAMT,calcium transport ATPase, andplasma membrane ATPasewere also discovered participating in GA signal transduction. Transcription factors includingWRKY7,NAC2,NAC11, andCBFspecially regulated phosphorylation and glycosylation during the ubiquitination degradation process ofDELLAproteins. These transcription factors also activated in abscisic acid metabolism. A large number of transcription factors such asWRKY21,WRKY22,NAC1,AP2,EREB1,P450, andCYP81that both regulate gibberellin signaling and low-temperature signals have also been found. Finally, the molecular mechanism of GA floral induction pathway in the LA hybrid lily ‘Aladdin’ was constructed.Keywords:GA floral induction pathway;LA hybrid lily;transcriptome RNA-Seq;GCN;qRT-PCR
研究领域:
植物学
·
遗传学
·
分子生物学
·
植物生理学
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DOI
PubMed
高影响因子
JOURNAL OF FISH BIOLOGY
·
2016
·Vol. 88
·期 5
·1693-1707
·
IF 1.246
W. T. Zhang,K. Liu,J. S. Xiang,L. Y. Zhang,W. J. Liu,Z. D. Dong,Y. Z. Li,H. L. Li,S. L. Chen,N. Wang
To elucidate the effect of thyroid hormone receptor α A ( thraa ) on metamorphosis, the full length cDNA of half-smooth tongue sole Cynoglossus semilaevis was cloned. The relative gene transcript level of thraa at different development stages was quantified using real-time PCR. Transcription of thraa increased and declined rapidly during metamorphosis. Hyperthyroidism was induced in juveniles and larvae with exposure to T3 and T4, and hypothyroidism with thiourea (TU), 2-mercapto-1-methylimidazole (MMI). thraa mRNA was higher in fish treated for 6 days with MMI than in untreated controls, although inhibited larvae did not complete metamorphosis. The addition of exogenous T4 reversed this effect in the MMI-treated group, but not in the TU-treated group. In situ hybridization revealed progressive tail end of body growth and change during developmental stages, with corresponding changes in thraa expression. This process may be induced by thyroid hormones with thraa as a major mediator. The morphological changes of tip of the tail may be associated with the development of lateral swimming.
研究领域:
分子生物学
·
内分泌学
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发育生物学
·
水产养殖
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DOI
PubMed
高影响因子
BIOMEDICINE & PHARMACOTHERAPY
·
2022
·
IF 7.419
Kaifang Yi,Shuobo Liu,Peiyi Liu,Xingwei Luo,Jinfeng Zhao,Fengbin Yan,Yushan Pan,Jianhua Liu,Yajun Zhai,Gongzheng Hu
It has been recognized that colistin resistance is a growing problem that seriously impairs the clinical efficacy of colistin against bacterial infections. One strategy that has been proven to have therapeutic effect is to overcome the widespread emergence of antibiotic-resistant pathogens by combining existing antibiotics with promising non-antibiotic agents. In this work, antibiotic susceptibility testing, checkerboard assays and time-kill curves were used to investigate the antibacterial activity of the individual drugs and the potential synergistic activity of the combination. The molecular mechanisms of tetrandrine in combination with colistin were analyzed using fluorometric assay and Real-time PCR. To predict possible interactions between tetrandrine and MCR-1, molecular docking assay was taken. Finally, we evaluated the in vivo efficacy of tetrandrine in combination with colistin against MCR-positive Salmonella . Overall, the combination of tetrandrine and colistin showed significant synergistic activity. In-depth mechanistic analysis showed that the combination of tetrandrine with colistin enhances the membrane-damaging ability of colistin, undermines the functions of proton motive force (PMF) and efflux pumps in MCR-positive bacteria. The results of molecular docking and RT-PCR analyses showed that tetrandrine not only affects the expression of mcr -1 but is also an effective MCR-1 inhibitor. Compared with colistin monotherapy, the combination of tetrandrine with colistin significantly reduced the bacterial load in vivo. Our findings demonstrated that tetrandrine serves as a potential colistin adjuvant against MCR-positive Salmonella .
研究领域:
微生物学
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药理学
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传染病学
·
分子生物学
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DOI
PubMed
高影响因子
MEDICAL MICROBIOLOGY AND IMMUNOLOGY
·
2025
·Vol. 214
·期 1
·1-13
·
IF 3
Ding Zhi-Wen,Xu Kai-Zhong,Dar Owias Iqbal,Yin Lu-Jun,Wang Ying-Jie,Liao Yun-tong,Wang Peng,Jia Ai-Qun
Burkholderia cenocepacia , an opportunistic pathogen, poses a significant threat to human health, necessitating the discovery of effective quorum sensing inhibitors (QSIs). In this study, the quorum sensing inhibitory effects of deferiprone (DFP) on the B. cenocepacia 162,638 were validated. Notably, DFP demonstrated an ability to inhibit and disrupt bacterial biofilms, reducing biofilm formation by 44.59% at 1/4 MIC (minimum inhibitory concentration) and 24.32% at 1/8 MIC concentrations. The study also investigated DFP’s impact on motility, virulence, and QS signal levels. LC-MS/MS analysis showed a gradual reduction in the QS molecule C6-HSL as DFP concentrations increased. Additionally, DFP’s non-hemolytic properties and safety profile, as verified in Galleria mellonella infection models, highlighted its biocompatibility. RT-qPCR results further indicated that DFP downregulated QS-related gene expression, particularly those involved in ferric uptake regulation protein (Fur). Molecular docking studies identified Fur as a key target for DFP’s inhibitory action. Collectively, DFP was shown as a potential QSI with practical applications for controlling B. cenocepacia infections.
研究领域:
微生物学
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传染病学
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药理学
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分子生物学
✓ 已验证
产品关联
DOI
PubMed
高影响因子
Analytical Methods
·
2023
·Vol. 15
·期 26
·3240-3250
·
IF 3.1
Ruihua Tang,Xueyan Yan,Min Li,Aoqi Du,Hui Yang,Huancai Yin,Mingyue Xie
Nucleic acid detection technologies have been widely utilized for various diseases. Conventional laboratory tests are less suitable for use in resource-limited settings as they are time-consuming, high-cost, complex, and heavily dependent on benchtop equipment. Rapid nucleic acid detection methods that consist of rapid nucleic acid extraction steps could overcome these challenges. A paper-based platform has been utilized to develop various rapid nucleic acid extraction methods owing to its cost-effectiveness, portability, and easy-modification. However, the existing paper-based nucleic acid extraction technologies mainly focus on improving the adsorption capacity of nucleic acids without reducing the non-specific adsorption capacity of proteins. In this study, paper-based nucleic acid extraction technology with wash-free, elution-free, and low protein adsorption was developed. The fabrication of paper involves the mixing of polyethylene glycol (PEG)-modified cotton fiber, chitosan (COS)-modified cotton fiber, and cotton fiber to form PEG-modified cotton fiber/chitosan-modified cotton fiber/cotton fiber (PEG-CF/COS-CF/CF) paper by the wet molding method. The result showed that PEG-CF/COS-CF/CF paper has a desirable pore size (23.9 ± 4.03 μm), good mechanical strength (dry: 9.37 Mpa and wet: 0.28 Mpa), and hydrophilicity (contact angle: 42.6° ± 0.36°). NH3+ groups of COS and OH− groups of PEG were observed on its surface and the adsorption efficiency of nucleic acid in TE buffer was 42.48% ± 0.30%. The limit of detection of pure DNA with this PEG-CF/COS-CF/CF paper by qPCR was as low as 25 ng. Additionally, this platform could successfully extract nucleic acid from 30 μL of a saliva sample, highlighting its potential use for clinical sample testing. The proposed paper-based nucleic acid extraction platform shows tremendous potential for disease diagnosis in resource-limited settings.
研究领域:
生物技术
·
分子生物学
·
诊断技术
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材料科学
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产品关联
DOI
PubMed
高影响因子
BIOSENSORS & BIOELECTRONICS
·
2016
·
IF 7.476
Yingshu Guo,Yujie Wang,Guangxu Yang,Jing-Juan Xu,Hong-Yuan Chen
Here, a mass-sensitive microRNA sensing surface is developed by utilizing a probe DNA/gold nanoparticles (GNP)/dendrimer composite coupling with an enzymatic amplification process. The probe DNA/GNP/dendrimer composite is prepared via the covalent coupling between the NH 2 groups in PAMAM or DNA and the COOH group on GNP. Target microRNA binds to a stem-loop-structured DNA on maganatic NPs, forming a heteroduplex . By enzyme recycling amplification, a large number of linker sequences are produced on MNPs . Via the combination of probe DNA , the linker DNA on MNPs and a capture DNA on gold chip, the DNA/GNP/dendrimer probe could be assembled on the surface of gold chip inducing a sensitive frequency response. It presents an excellent performance in microRNA-203 quantification with a detection linear range of 1.0×10 −11 –1.0×10 −9 M. Both the success in discriminating expressing miroRNA-203 in MCF-7 cell and the well agreement with the commercial qRT-PCR detection method implied its potential application in early cancer diagnosis for future.
研究领域:
生物传感器
·
纳米技术
·
分子生物学
·
癌症诊断
✓ 已验证
DOI
PubMed
高影响因子
危险品化学品经营许可证（带存储）