A real time S1 assay at neutral pH based on graphene oxide quenched fluorescence probe

AbstractAs the extracellular nuclease of Aspergillus, S1 nuclease can split single and double-stranded DNA into oligo- or mononucleotides, while preferentially digests single-stranded nucleic acids. Furthermore, the existence of S1 can be the standard to identify Aspergillus and used to evaluate the severity of Aspergillosis. Herein, a simple and sensitive fluorescent sensing platform for S1 assay was developed based on the S1-induced DNA strand scission and the difference in affinity of graphene oxide (GO) for single-stranded DNA containing different bases. This platform was applied to monitor S1 activity and study the kinetics in real time. Results indicated that the detection limit is 0.5U/mL. The Km and kcat at 45°C, are 1.4±0.12μM and 0.6min−1, respectively. Moreover, by monitoring the effect of chemical drugs on S1 activity, we found that 2mM of erythromycin, sodium penicillin, carbenicillin disodium and ampicillin can inhibit S1 activity about 8%, 60%, 61% and 66%, respectively, while gentamycin sulfate is a stimulator. Overall, the assay platform based on graphene oxide quenched fluorescence probe is successfully constructed to study the enzymatic activity of S1 and used for screening antibiotics

Folate monoglutamate in cereal grains: Evaluation of extraction techniques and determination by LC-MS/MS

Folates are essential micronutrients for human health. To determine the total folate content, the extraction and quantification of seven monoglutamate folate derivatives in cereals (maize, rice, and wheat) were optimised and validated in this study. Di-enzyme treatment with α-amylase and rat conjugase was proved ideal for folate extraction from the cereal grains. The quantification method by liquid chromatography-tandem mass spectrometry was validated based on its matrix effect, linearity, sensitivity, recovery, inter-day and intra-day precision. The limits of detection and quantification of folate derivatives ranged from 0.03–0.88 and 0.1–1.0 μg/100 g among the three cereal samples. The absolute recoveries of most folate derivatives were 72–96 % for these cereal samples, with the exception of dihydrofolate, tetrahydrofolate, and folic acid (44–65 %). The inter-day and intra-day precisions were < 12 % for the three cereals. Analysis of folate content and composition in several cereal grains showed that the total folate levels were approximately 26−37 μg/100 g, with 5-methyl-tetrahydrofolate and 5-methyl-tetrahydrofolate as the dominant. MeFox, an oxidation product of 5-methyltetrahydrofolate, was detected at concentrations 20–39-fold higher than those of total folates in rice and wheat grains. This validated method is an efficient approach for folate determination in cereal crops

Involvement of Fenton chemistry in rice straw degradation by the lignocellulolytic bacterium Pantoea ananatis Sd-1

Additional file 7: Table S2. Primers used for quantitative real time-PCR

Genome-wide identification and transcriptional analysis of folate metabolism-related genes in maize kernels

BACKGROUND: Maize is a major staple food crop globally and contains various concentrations of vitamins. Folates are essential water-soluble B-vitamins that play an important role as one-carbon (C1) donors and acceptors in organisms. To gain an understanding of folate metabolism in maize, we performed an intensive in silico analysis to screen for genes involved in folate metabolism using publicly available databases, followed by examination of the transcript expression patterns and profiling of the folate derivatives in the kernels of two maize inbred lines. RESULTS: A total of 36 candidate genes corresponding to 16 folate metabolism-related enzymes were identified. The maize genome contains all the enzymes required for folate and C1 metabolism, characterized by highly conserved functional domains across all the other species investigated. Phylogenetic analysis revealed that these enzymes in maize are conserved throughout evolution and have a high level of similarity with those in sorghum and millet. The LC-MS analyses of two maize inbred lines demonstrated that 5-methyltetrahydrofolate was the major form of folate derivative in young seeds, while 5-formyltetrahydrofolate in mature seeds. Most of the genes involved in folate and C1 metabolism exhibited similar transcriptional expression patterns between these two maize lines, with the highest transcript abundance detected on day after pollination (DAP) 6 and the decreased transcript abundance on DAP 12 and 18. Compared with the seeds on DAP 30, 5-methyltetrahydrofolate was decreased and 5-formyltetrahydrofolate was increased sharply in the mature dry seeds. CONCLUSIONS: The enzymes involved in folate and C1 metabolism are conserved between maize and other plant species. Folate and C1 metabolism is active in young developing maize seeds at transcriptional levels

Manipulation of Metabolic Pathways to Develop Vitamin-Enriched Crops for Human Health.

Vitamin deficiencies are major forms of micronutrient deficiencies, and are associated with huge economic losses as well as severe physical and intellectual damages to humans. Much evidence has demonstrated that biofortification plays an important role in combating vitamin deficiencies due to its economical and effective delivery of nutrients to populations in need. Biofortification enables food plants to be enriched with vitamins through conventional breeding and/or biotechnology. Here, we focus on the progress in the manipulation of the vitamin metabolism, an essential part of biofortification, by the genetic modification or by the marker-assisted selection to understand mechanisms underlying metabolic improvement in food plants. We also propose to integrate new breeding technologies with metabolic pathway modification to facilitate biofortification in food plants and, thereby, to benefit human health

Adsorbent-Assisted In Situ Electrocatalysis: Highly Sensitive and Stable Electrochemical Sensor Based on AuNF/COF-SH/CNT Nanocomposites for the Determination of Trace Cu(II)

The weak conductivity of covalent organic frameworks (COFs) limits their wide application in electrochemical sensors. Here, a novel electrochemical sensor (AuNFs/COF-SH/ CNTs/GCE) was designed and constructed by a one-step electrochemical deposition of Au nanoflowers (AuNFs) on a hybrid nanocomposite of sulfhydryl-functional covalent organic framework/carbon nanotubes (COF-SH/CNTs) for the determination of Cu(II) in seawater. AuNFs/COF-SH/CNTs/GCE was prepared in a three-step method including in situ synthesis, post synthesis, and one-step electrochemical deposition. The adsorption and catalytic performances of the modified electrode were improved based on the mechanism of ???adsorption-catalysis stripping determination???. By combining the excellent catalytic properties of the AuNFs with the good adsorption capacity of COF-SH toward Cu(II), as well as the good conductivity of CNTs, the AuNFs/COF-SH/CNTs/GCE exhibited excellent performance for the determination of Cu(II). Through optimization of the experimental conditions, a low detection limit of 0.47 nM and a wide linear range of 1.6 nM-4.7 ??M were obtained. Moreover, the sensor possessed good stability with a relative standard deviation of less than 5% after 20 repeated measurements. The results are consistent with certified values when used for the determination of certified reference materials. In summary, the AuNFs/COF-SH/CNTs/GCE, with high stability and sensitivity, has been successfully applied for the determination of Cu(II) in seawater samples with satisfactory results

适体标记银纳米团簇用于细胞图像和体外粘蛋白1检测。

Development of specific cell imaging technology for accurate tumor early diagnosis and evaluation of drug therapeutic efficiency is in great demand. In this study, a simple and sensitive fluorescence method for Mucin1 (MUC1) image in situ and quantitative assay in vitro has been established using APT-tagged silver nanoclusters (APT-Agnes) containing a recognition unit of MUC1 aptamer as the label-free fluorescence probe. The principle of the method is that specific recognition and binding of MUC1 with aptamer can result in the fluorescence quenching of APT-Agnes. The method for MUC1 assay showed a linear range from 0.1 to 100 NM with a limit of detection of 0.05 nM. Furthermore, the fluorescent probe of APT-AgNCs was successfully used for detection of MUC1 in serum and MCF-7 cell imaging. In our point, the above results demonstrated that the new simple method provided an alternative for direct quantitative assay of MUC1 in homogeneous solution and cell imaging, which is helpful for biomedical study and clinical diagnosis related with MUC1

Detection of Cu2+ in Water Based on Histidine-Gold Labeled Multiwalled Carbon Nanotube Electrochemical Sensor

Based on the strong interaction between histidine and copper ions and the signal enhancement effect of gold-labeling carbon nanotubes, an electrochemical sensor is established and used to measure copper ions in river water. In this study the results show that the concentrations of copper ion have well linear relationship with the peak current in the range of 10−11–10−7 mol/L, and the limit of detection is 10−12 mol/L. When using this method to detect copper ions in the Xiangjiang River, the test results are consistent with the atomic absorption method. This study shows that the sensor is convenient to be used in daily monitoring of copper ions in river water

Nafion assisted preparation of prussian blue nanoparticles and its application in electrochemical analysis of L-ascorbic acid

Due to the high sensitivity, simplicity and low cost, electrochemistry methods based on nanoparticles have been considered as an attractive technique for L-ascorbic acid. Herein, prussian blue nanoparticles (PBNPs) were synthesized by a fast and convenient synthesis strategy with Nafion and K4[Fe (CN)(6)] 3H2O. The PBNPs have uniform grain size of about 11.9 nm. PBNPs modified glassy carbon electrode (GCE) was used to construct electrochemical sensor for L-ascorbic acid (AA) detection and the optimization condition was studied. The electrochemical results indicate that PBNPs/GCE sensor shows a linear response to L-ascorbic acid in the range of 1-1100 mu M with a detection limit of 0.47 mu M and sensitivity of 22.9 mu A.mM(-1).cm(-2) at a working potential of 0.24 V (vs. SCE). The stability and reproducibility of the same PBNPs/GCE are RSD = 0.7% and RSD = 1.3%, respectively. The excellent characteristics of this sensor demonstrates it is a useful tool for accurate determination of L-ascorbic acid in medicine samples such as a vitamin C tablet and for Vitamin C chewable tablets

Comparative Transcriptome Analysis Reveals Mechanisms of Folate Accumulation in Maize Grains.

Previously, the complexity of folate accumulation in the early stages of maize kernel development has been reported, but the mechanisms of folate accumulation are unclear. Two maize inbred lines, DAN3130 and JI63, with different patterns of folate accumulation and different total folate contents in mature kernels were used to investigate the transcriptional regulation of folate metabolism during late stages of kernel formation by comparative transcriptome analysis. The folate accumulation during DAP 24 to mature kernels could be controlled by circumjacent pathways of folate biosynthesis, such as pyruvate metabolism, glutamate metabolism, and serine/glycine metabolism. In addition, the folate variation between these two inbred lines was related to those genes among folate metabolism, such as genes in the pteridine branch, para-aminobenzoate branch, serine/tetrahydrofolate (THF)/5-methyltetrahydrofolate cycle, and the conversion of THF monoglutamate to THF polyglutamate. The findings provided insight into folate accumulation mechanisms during maize kernel formation to promote folate biofortification