Bin Qi

Building An Aptamer/Graphene Oxide FRET Biosensor for One-Step Detection of Bisphenol A

Bisphenol A (BPA) is an important industrial chemical for polycarbonate (PC) and epoxy resins in paper and plastic industries. In our work, a kind of new method for detection of BPA was designed based on graphene oxide and anti-BPA aptamer. The graphene oxide can specifically adsorb and quench the fluorescence of fluorescently modified ssDNA probes. Meanwhile, the BPA can combine with anti-BPA optamer and switch its configuration to prevent the aptamer from adsorbing on the surface of graphene oxide (GO). Under different concentrations of BPA, based on the target-induced conformational change of anti-BPA aptamer and the interactions between the fluorescently modified anti-BPA aptamer (FAM-ssDNA) and GO, the experimental results show that the intensity of the fluorescence signal was changed. A low limit of detection of 0.05 ng/mL was obtained in the range 0.1-10 ng/mL. In addition, the specificity was outstanding among analogues of BPA. The recovery rate in actual water samples spiked with BPA can be 96.0% to 104.5%. The developed method was successfully used to determine BPA in actual water samples.

Highly sensitive colorimetric sensor for Hg2+ detection based on cationic polymer/DNA interaction

The detection of ultralow concentrations of mercury is a currently significant challenge. Here, a novel strategy is proposed: the colorimetric detection of Hg(2+) based on the aggregation of gold nanoparticles (AuNPs) driven by a cationic polymer. In this three-component system, DNA combines electrostatically with phthalic diglycol diacrylate (PDDA) in a solution of AuNPs. In the presence of Hg(2+), thymine (T)-Hg(2+)-T induced hairpin turns are formed in the DNA strands, which then do not interact with PDDA, enabling the freed PDDA to subsequently facilitate aggregation of the AuNPs. Thus, according to the change in color from wine-red to blue-purple upon AuNPs aggregation, a colorimetric sensor is established to detect Hg(2+). Under optimal conditions, the color change is clearly seen with the naked eye. A linear range of 0.25-500nM was obtained by absorption spectroscopy with a detection limit of approximately 0.15nM. Additionally, the proposed method shows high selectivity toward Hg(2+) in the presence of other heavy metal ions. Real sample analysis was evaluated with the use of lake water and the results suggest good potential for practical application.

Ultrasensitive detection of lead ions based on a DNA-labelled DNAzyme sensor

Here, we report a facile approach for highly sensitive and selective detection of aqueous lead ions that uses a real-time quantitative polymerase chain reaction technology and a lead-dependent DNAzyme, termed GR-5. In this method, the substrate DNA is cleaved at the site of the adenosine ribonucleotide by GR-5 DNAzyme in the presence of lead ions, resulting in a decrease in template DNA available for PCR and a consequent change in signal detection (cycle threshold (Ct) value). This novel approach takes advantage of the exponential amplification of PCR and the specific recognition of the GR-5 lead-dependent DNAzyme to provide Pb2+-specific detection with an excellent linear relationship between Ct value and Pb2+ concentration within a range of 1–500 nM. The correlation coefficient of the standard curve was 0.9898, and the limit of detection was 0.7 nM. Moreover, this sensor showed good selectivity for Pb2+ ions over other metal ions.

Mercury–DNA interaction based detection of mercury ions by DNA amplification with high sensitivity and selectivity

An effective and facile approach for mercury detection is essential for environmental protection and human health. In this study, we propose a novel strategy to develop a highly sensitive and selective method for the detection of mercury ions using real-time quantitative polymerase chain reaction (RT-qPCR) technology and the specific binding of Hg2+ in thymine–thymine mismatch complexes. The signal for detection of Hg2+ was generated using RT-qPCR amplification of a template strand of DNA. The sensor exhibited an excellent linear response between the cycle threshold (Ct) and mercury ion concentration in the range of 0.05–100 nM. Under optimised conditions, the detection limit of this method for aqueous Hg2+ was as low as 30 pM. In addition, this system showed good selectivity for Hg2+ over other metal ions. This new approach shows significant potential for detection of Hg2+ in water and other samples.

Endogenic oxidative stress response contributes to glutathione over-accumulation in mutant Saccharomyces cerevisiae Y518.

Mechanisms of glutathione (GSH) over-accumulation in mutant Saccharomyces cerevisiae Y518 screened by ultraviolet and nitrosoguanidine-induced random mutagenesis were studied. Y518 accumulated higher levels of GSH and l-cysteine than its wild-type strain. RNA-Seq and pathway enrichment analysis indicated a difference in the expression of key genes involved in cysteine production, the GSH biosynthesis pathway, and antioxidation processes. GSH1, MET17, CYS4, GPX2, CTT1, TRX2, and SOD1 and the transcriptional activators SKN7 and YAP1 were up-regulated in the mutant. Moreover, Y518 showed a dysfunctional respiratory chain resulting from dramatically weakened activity of complex III and significant elevation of intracellular reactive oxygen species (ROS) levels. The supplementation of antimycin A in the culture of the parent strain showed equivalent changes of ROS and GSH level. This study indicates that defective complex III prompts abundant endogenic ROS generation, which triggers an oxidative stress response and upregulation of gene expression associated with GSH biosynthesis. This finding may be helpful for developing new strategies for GSH fermentation process optimization or metabolic engineering.

Biosynthesis of (R)-2-hydroxy-3-phenylpropionic acid using whole recombinant Escherichia coli cells in an aqueous/n-octane biphasic system

Abstract(R)-2-hydroxy-3-phenylpropionic acid (PLA) is an ideal antimicrobial compound with broad-spectrum activity against a wide range of Gram-positive bacteria, some Gram-negative bacteria, and fungi. We studied the bioconversion of phenylpyruvate (PPA) to PLA using whole recombinant Escherichia coli cells in a series of buffer/organic solvent systems. Octane was found to be the best organic solvent. The optimum volume ratio of the water phase to the n-octane phase, conversion temperature, substrate concentration, and cell concentration were 6:4, 40 °C, 12.5 g/L, and 30 g/L wet cells, respectively. Under the optimized conditions, the average PLA productivity in the aqueous/n-octane system was 30.69% higher than that in the aqueous system, and 32.31 g/L PLA was obtained with the use of a stirred reactor (2-L scale). Taken together, our findings indicated that PLA biosynthesis was more efficient in an aqueous/n-octane biphasic system than in a monophasic aqueous system. The proposed biphasic system is an effective strategy for enhancing PLA yield and the biosynthesis of its analogues.概 要目 的通过构建水/正辛烷双相体系用于苯基乳酸的合成, 有效减轻产物抑制作用, 改善细胞活力, 增加苯基乳酸的产率及转化率。创新点在苯基乳酸的全细胞合成中, 苯基乳酸的抑制作用导致微生物细胞活力降低, 产率较低。 本研究旨在降低产物抑制作用来增加苯基乳酸的产率及转化率。方 法从极性不同的几种有机溶剂中筛选能够增加苯基乳酸产量的有机溶剂与水形成双相体系, 在水/正辛烷双相体系中利用重组大肠杆菌全细胞转化法合成苯基乳酸, 并对该体系转化过程进行优化。 部分产物转移至正辛烷相中, 减轻了产物抑制作用。结 论本研究通过构建水/正辛烷双相体系全细胞合成苯基乳酸, 优化了转化条件。 正辛烷相的存在减轻了产物抑制作用。 在最佳条件下, 双相体系分批补料合成的苯基乳酸明显高于纯水相系统的产量。

Enantioselective Biosynthesis of l-Phenyllactic Acid by Whole Cells of Recombinant Escherichia coli

Background: l-Phenyllactic acid (l-PLA)—a valuable building block in the pharmaceutical and chemical industry—has recently emerged as an important monomer in the composition of the novel degradable biocompatible material of polyphenyllactic acid. However, both normally chemically synthesized and naturally occurring phenyllactic acid are racemic, and the product yields of reported l-PLA synthesis processes remain unsatisfactory. Methods: We developed a novel recombinant Escherichia coli strain, co-expressing l-lactate dehydrogenase (l-LDH) from Lactobacillus plantarum subsp. plantarum and glucose dehydrogenase (GDH) from Bacillus megaterium, to construct a recombinant oxidation/reduction cycle for whole-cell biotransformation of phenylpyruvic acid (PPA) into chiral l-PLA in an enantioselective and continuous manner. Results: During fed-batch bioconversion with intermittent PPA feeding, l-PLA yield reached 103.8 mM, with an excellent enantiomeric excess of 99.7%. The productivity of l-PLA was as high as 5.2 mM·h−1 per OD600 (optical density at 600 nm) of whole cells. These results demonstrate the efficient production of l-PLA by the one-pot biotransformation system. Therefore, this stereoselective biocatalytic process might be a promising alternative for l-PLA production.

Microbial Community Structure and Diversity of Shrimp Paste at Different Fermentation Stages

High-throughput sequencing was used to reveal the highly diverse bacterial populations in shrimp paste at different fermentation stages. We studied three stages of fermentation and obtained 448,916 reads. Using this approach, we revealed the presence of 30 phyla, 55 classes, 86 orders, 206 families and 695 genera of bacteria in the shrimp paste. Shrimp paste in fermentation metaphase had a more diverse microbiota than that in fermentation prophase and fermentation anaphase. Diversity appeared greatest in fermentation anaphase. The four dominant phyla were Proteobacteria, Firmicutes, Actinobacteria, and Bacteroidetes. The most common genera were Psychrobacter, Halomonas, Bacillus, Alteribacillus, and Lactococcus. Their content varied at different stages of fermentation. All the microbiome presented a variety of changes in the microbial diversity of shrimp paste. Importance Most research on the microbial diversity of shrimp paste has focused on the shrimp culture environment, or the chemical composition and sensory attributes of the paste. Little research has been conducted on the microbial diversity and composition of shrimp paste. The relationship between microbes and the flavor and quality of shrimp paste has thus been unknown. We therefore analyzed the microbial composition and variation of shrimp paste at different stages of fermentation. The dominant bacteria in fermentation prophase, metaphase, and anaphase were identified. Our preliminary findings give some insight into which microbes contribute to the flavor of shrimp paste and suggest how to improve its flavor. In addition, our findings are relevant to optimizing the production of shrimp paste and guaranteeing its quality and safety.

Effect of loach paste on the liver and immune organs of D-galactose-induced ageing mice

ABSTRACT We examined the effects of loach paste (LP) on liver and immune organs of d-galactose (DG)-induced aging mice. The results showed that LP can alleviate the atrophy of spleen and thymus, and restore the blood glucose levels. LP was particularly beneficial to liver. Liver indices and activities of aspartate aminotransferase, alanine aminotransferase, and alkaline phosphatase reduced with high doses of LP. Moreover, LP can improve the antioxidant system in liver by increasing the activities of total superoxide dismutase, glutathione peroxidase, and catalase, reducing malondialdehyde concentration. Furthermore, LP can improve immunity and reduce the inflammation of liver. The swelling of liver cells was alleviated and the cell arrangement returned to normal in the LP group. In addition, gene expressions of HSP70 and tumour necrosis factor-α (TNF-α) in liver which can promote inflammatory cytokine decreased significantly. Thus, regular LP intake effectively relieved DG-induced oxidative stress and alleviated liver and immune organs damage.