Histidine modified Fe3O4 nanoparticles improving the ethanol yield and tolerance of Saccharomyces cerevisiae.

Saccharomyces cerevisiae, the primary microorganism involved in ethanol production, is hindered by the accumulation of ethanol, leading to reduced ethanol production. In this study, we employed histidine-modified Fe3O4 nanoparticles (His-Fe3O4) for the first time, to the best of our knowledge, as a method to enhance ethanol yield during the S. cerevisiae fermentation process. The results demonstrated that exposing S. cerevisiae cells to Fe3O4 nanoparticles (Fe3O4 NPs) led to increased cell proliferation and glucose consumption. Moreover, the introduction of His-Fe3O4 significantly boosted ethanol content by 17.3% (p < 0.05) during fermentation. Subsequent findings indicated that the increase in ethanol content was associated with enhanced ethanol tolerance and improved electron transport efficiency. This study provided evidence for the positive effects of His-Fe3O4 on S. cerevisiae cells and proposed a straightforward approach to enhance ethanol production in S. cerevisiae fermentation. The mediation of improved ethanol tolerance offers significant potential in the fermentation and bioenergy sectors.

A small-scale external quality assessment for PCR detection of group B streptococcus in China.

BACKGROUND: Group B streptococcus (GBS) is considered a leading cause of maternal and infant morbidity and mortality. Molecular diagnosis is a routinely used approach for GBS screening to protect pregnant women and prevent early-onset GBS neonatal disease. The objective of this study was to identify issues and guarantee the dependability of GBS molecular diagnosis by an external quality assessment (EQA) scheme. METHODS: The EQA panel comprised eight samples spiked with 10-fold dilutions of GBS suspension (20-2,000,000 copies/mL), and 2 negative control samples. The panels were coded randomly and distributed to participating laboratories for GBS detection. RESULTS: In total, 44 participating laboratories submitted results with eight commercial GBS PCR assays and one in-house assay. Among them, 36 obtained an acceptable or higher performance score, while 8 required improvement. Among the 440 results returned, 62 (14.1 %) were incorrect, including 5 false positives and 57 false negatives. CONCLUSIONS: Our small-scale EQA showed that most participating laboratories have reliable diagnostic capacities for GBS PCR detection. Nonetheless, further improvements in the detection performance of some laboratories are required, particularly with low-concentration samples. Our survey also reinforces the use of EQA as an essential tool to evaluate the overall proficiency of clinical laboratories.

Synthesis of N,Si co-doped carbon dots to establish a fluorescent sensor for Hg(II) detection with triple signal output.

Mercury is a heavy metal with extreme toxicity. Thus, it is of significance to develop an effective method for mercury ion detection with high performance. In this study, carbon dots doped with nitrogen and silicon (N,Si/CQDs) were successfully prepared from folic acid and N-[3-(trimethoxysily)propyl]-ethylenediamine. The N,Si/CQDs show an obvious cyan fluorescence of 460 nm with the radiation of 350 nm. The existence of mercury ions induces the fluorescence quenching of N,Si/CQDs due to photoinduced electron transfer, which was applied for the sensitive sensing of Hg(II). More importantly, the practical application of the N,Si/CQD probe was confirmed by measurements of Hg(II) in real samples of lake water, sorghum and rice. In addition, the N,Si/CQD nanoprobe was integrated on a sensing strip for specific detection of Hg(II). Quantitative measurement of Hg(II) was realized by the outstanding linearity between the diameter (or fluorescence intensity) of the fluorescence quenching ring and the concentration of mercury ions. The sensor shows potential for rapid detection with a triple signal readout on-site and represents a larger step towards practical applications.

Synthesis of two nitrogen-doped carbon quantum dots to construct fluorescence probes for sensitive Hg2+ detection with dual signal output.

The rapid and sensitive detection of heavy metal ions is of great importance in food safety and for the environment. Therefore, two novel probes, M-CQDs and P-CQDs, based on carbon quantum dots were utilized to detect Hg2+ based on fluorescence resonance energy transfer and photoinduced electron transfer mechanisms. The M-CQDs were prepared from folic acid and m-phenylenediamine (mPDA) using a hydrothermal method. Similarly, the novel P-CQDs were obtained according to the same synthetic procedure used to create M-CQDs except the mPDA was replaced with p-phenylenediamine (pPDA). Upon the addition of Hg2+ to the M-CQDs probe, the fluorescence intensity reduced significantly with a linear concentration range between 5 and 200 nM. The limit of detection (LOD) was calculated to be 2.15 nM. On the contrary, the fluorescence intensity of the P-CQDs was enhanced greatly after the addition of Hg2+. The Hg2+ detection was realized with a wide linear range from 100 to 5000 nM and the LOD was calculated to be as low as 52.5 nM. The fluorescence "quenching" and "enhancing" effect exhibited by the M-CQDs and P-CQDs, respectively, is due to the different distribution of -NH2 in the mPDA and pPDA precursors. Notably, paper-based chips modified with M/P-CQDs were established for visual Hg2+ sensing, demonstrating the possibility for real-time detection of Hg2+. Moreover, the practicality of this system was confirmed through the successful measurement of Hg2+ in tap water and river water samples.

Discrimination of Chinese green tea according to tea polyphenols using fluorescence sensor array based on Tb (III) and Eu (III) doped Zr (IV) metal-organic frameworks.

A facile and rapid fluorescence sensor array based on Tb (III) and Eu (III) doped Zr (IV) metal-organic frameworks was proposed for Chinese green tea discrimination. According to large porosity of Tb@UiO-66-(COOH)2 and Eu@UiO-66-(COOH)2, phenolic hydroxyl groups of tea polyphenols could coordinate with free carboxylic acid groups and was captured into the pores, which led to the disturbance of electronic structure of ligand and inhibited the energy transfer efficiency from ligand to Tb (III) and Eu (III) center, causing the fluorescence quenching effect. Based on Hierarchy Cluster Analysis and Linear Discrimination Analysis, the fluorescence sensor array was employed for successful tea polyphenols classification through the analysis of different fluorescence quenching effect to tea polyphenols. Green tea samples within different categories and grades were also successfully discriminated using this assay according to tea polyphenols, providing a new method for Chinese green tea identification.

Naked-eye detection of site-specific ssRNA and ssDNA using PAMmer-assisted CRISPR/Cas9 coupling with exponential amplification reaction.

Accurate and effective detection of single-stranded nucleic acids is vital in both disease diagnosis and pathological studies. Hence, we develop a PAMmer-assisted CRISPR/Cas9 system mediated G4-EXPAR (Cas-G4EX) strategy for site-specific detection of ssRNA and ssDNA. PAMmer-assisted CRISPR/Cas9 executes the site-specific cleavage of target ssRNA or ssDNA and released product fragment with the desired sequence at the 3'-terminal. This fragment serves as a primer to activate subsequent sequence-dependent exponential amplification reaction (EXPAR). The G-rich EXPAR products assembles with hemin to form a G-Quadruplex (G4/hemin). G4/hemin catalyzes ABTS-H2O2 system with the appearance of vivid green color, realizing naked-eye analysis. Cas-G4EX integrates the superiority of CRISPR/Cas9 and EXPAR, presenting outstanding site-specific recognition and high-performance amplification efficiency. Meanwhile, the programmability of CRISPR/Cas9 system makes the proposed method become a universal detection paradigm for any ssRNA or ssDNA. Cas-G4EX assay shows the linear relationship from 250 aM to 2.5 nM for ssRNA detection with the actual LOD of 250 aM, and that ranges from 100 aM to 1 nM for ssDNA detection with the actual LOD of 100 aM. Additionally, the acceptable recoveries of 101.48%-109.61% for ssRNA and 93.25%-111.98% for ssDNA in real detection of human serum are obtained for detection of single-strand nucleic acid in real samples. Cas-G4EX also exhibits the excellent discrimination for single-base mutation of single-stranded nucleic acids. Therefore, Cas-G4EX assay provides a promising platform in the applications of molecular diagnosis and pathological analysis.

Dual Methylation-Sensitive Restriction Endonucleases Coupling with an RPA-Assisted CRISPR/Cas13a System (DESCS) for Highly Sensitive Analysis of DNA Methylation and Its Application for Point-of-Care Detection.

High-performance detection of DNA methylation possesses great significance for the diagnosis and therapy of cancer. Herein, for the first time, we present a digestion strategy based on dual methylation-sensitive restriction endonucleases coupling with a recombinase polymerase amplification (RPA)-assisted CRISPR/Cas13a system (DESCS) for accurate and sensitive determination of site-specific DNA methylation. This dual methylation-sensitive restriction endonuclease system selectively digests the unmethylated target but exhibits no response to methylated DNA. Therefore, the intact methylated DNA target triggers the RPA reaction for rapid signal amplification. In contrast, the digested unmethylated target initiates no RPA reaction. RPA products with a T7 promoter can execute the T7 transcription in the presence of T7 RNA polymerase to generate a large number of single-stranded RNA (ssRNA). This ssRNA can be recognized by CRISPR/Cas13a to induce the ssRNase activity of Cas13a, showing the indiscriminate cleavage of the collateral FQ reporter to release the fluorescence signal. With such a design, by combining the unique features of dual methylation-sensitive restriction endonucleases with RPA-assisted CRISPR/Cas13a, the DESCS system not only presents the rapid and powerful signal amplification for the determination of methylated DNA with ultrahigh sensitivity but also effectively eliminates the false positive influences from incomplete digestion of the unmethylated target. More importantly, 0.01% methylation level can be effectively distinguished with the existence of excess unmethylated DNA. In addition, the DESCS assay is integrated into the lateral flow biosensor (LFB) for the point-of-care determination of DNA methylation. In view of the superiorities in high sensitivity, outstanding selectivity, and ease of operation, the DESCS system will provide a reliable assay for site-specific analysis of methylation.

Target-induced transcription amplification to trigger the trans-cleavage activity of CRISPR/Cas13a (TITAC-Cas) for detection of alkaline phosphatase.

Herein, an ultra-sensitive alkaline phosphatase (ALP) sensing strategy is developed by target-induced transcription amplification to trigger the trans-cleavage activity of Cas13a (TITAC-Cas). A double-stranded DNA duplex integrating a T7 promoter with 5'-phosphate and a transcription template (5'P-dsDNA) serves as the ALP substrate. In the absence of ALP, 5'P-dsDNA can be degraded by the λexo, leading to the subsequent transcription failure. In the presence of ALP, dephosphorylation reaction converts the 5'P-dsDNA to 5'OH-dsDNA and provides the protection for T7 promoter against the λexo-digestion. The intact T7 promoter of 5'OH-dsDNA can activate T7 transcription to produce a mass of single-stranded RNA (ssRNA). The ssRNA products possess a full complementarity to the spacer of crRNA and activate the ssRNase activity of CRISPR/Cas13a. As a result, Cas13a exhibits the indiscriminate cleavage of collateral FQ-reporter to release significant fluorescence signal, realizing the ultra-sensitive detection of ALP. Due to the triple signal amplification (ALP self-catalysis, T7 transcription amplification, and trans-cleavage of CRISPR/Cas13a), TITAC-Cas assay shows the ultra-sensitive detection of ALP activity with a wide linear range from 0.008 to 250 U∙L-1). The LOD is calculated to be 6 ± 0.52 mU∙L-1. TITAC-Cas assay is also successfully applied for analysis of ALP activity in HepG2 cell lysate with high fidelity. In addition, this method is employed to screen ALP inhibitor.

Simultaneous measurement of Cr(III) and Cu(II) based on indicator-displacement assay using a colorimetric nanoprobe.

High-performance analysis of heavy metal ions is great importance in both environment and food safety. In this work, a facile and reliable colorimetric sensor was presented for simultaneous detection of Cu2+ and Cr3+ based on indicator-displacement assay (IDA). As a typical silicate nanomaterials, ZnSiO3 hollow nanosphere (ZSHS) exhibited an outstanding ion exchange capacity. Zincon was incorporated with the ZSHS to form a zincon/ZSHS hybrid ionophore with a blue color. Upon the addition of Cr3+, IDA reaction and selective ion exchange occurred with the color change of zincon/ZSHS ionophore from blue to yellow. With such a design, colorimetric measurement of Cr3+ was realized. The linear concentration for Cr3+ detection ranged from 0.5 µM to 75 µM with the LOD of 83.2 nM. Furthermore, we also screened different kinds of complexing agents that may respond with zincon/ZSHS ionophore and various metal ions. It was found that tartaric acid (TA) showed the chelation capability of Zn2+-TA is stronger than that of Zn2+-zincon. Thus zincon/ZSHS/TA presented a yellow color due to the chelation reaction of Zn2+-TA, releasing the zincon as a free state. After addition of Cu2+, a stronger chelation reaction of Cu2+-zincon occurred. This process involved in the color change from yellow to blue and realized colorimetric measurement of Cu2+. The detection limit of Cu2+ was calculated to be 43.7 nM with linear range from 0.1 to 20 µM. In addition, the zincon/ZSHS nanoprobe was successfully applied for simultaneous measurement of Cu2+ and Cr3+ in sorghum and river water, indicating that the zincon/ZSHS nanoprobe provided a promising sensing platform in environment and food safety.