Comparative Study of Ultrasonic Vibration-Assisted Die-Sinking Micro-Electrical Discharge Machining on Polycrystalline Diamond and Titanium.

Die-sinking micro-electrical discharge machining (micro-EDM) is a potential method used to fabricate intricate structures without complex electrode motion planning and compensation. However, machining efficiency and poor discharge states are still bottlenecks. This study conducted a comparative investigation into the impact of ultrasonic vibration on die-sinking micro-EDM of polycrystalline diamond (PCD) and pure titanium (TA2). By adjusting discharge parameters, this study systematically evaluated the influence of ultrasonic vibration on these two materials based on discharge waveforms, motion trajectories, effective discharge counts and groove profiles. At an open-circuit voltage of 100 V, ultrasonic vibration promotes die-sinking micro-EDM of PCD. However, when the open-circuit voltage increases to 200 V, ultrasonic vibration exhibits inhibitory effects in general. Conversely, for TA2, ultrasonic vibration shows a promoting effect at both voltages, indicating the differences of ultrasonic vibration-assisted die-sinking micro-EDM on PCD and TA2. For PCD, ultrasonic cavitation improves the discharge gap environment, accelerating the removal of discharge debris. For TA2, due to its poor thermal conductivity, ultrasonic cavitation acts to break the arc, accelerating heat transfer. These research findings provide guidance for ultrasonic vibration-assisted die-sinking micro-EDM in industrial applications.

Extended Enrichment for Ultrasensitive Detection of Low-Frequency Mutations by Long Blocker Displacement Amplification.

Detecting low-frequency DNA mutations hotspots cluster is critical for cancer diagnosis but remains challenging. Quantitative PCR (qPCR) is constrained by sensitivity, and allele-specific PCR is restricted by throughput. Here we develop a long blocker displacement amplification (LBDA) coupled with qPCR for ultrasensitive and multiplexed variants detection. By designing long blocker oligos to perfectly match wildtype sequences while mispairing with mutants, long blockers enable 14-44 nt enrichment regions which is 2-fold longer than normal BDA in the experiments. For wild template with a specific nucleotide, LBDA can detect different mutation types down to 0.5 % variant allele frequency (VAF) in one reaction, with median enrichment fold of 1,000 on 21 mutant DNA templates compared to the wild type. We applied LBDA-qPCR to detect KRAS and NRAS mutation hotspots, utilizing a single plex assay capable of covering 81 mutations and tested in synthetic templates and colorectal cancer tissue samples. Moreover, the mutation types were verified through Sanger sequencing, demonstrating concordance with results obtained from next generation sequencing. Overall, LBDA-qPCR provides a simple yet ultrasensitive approach for multiplexed detection of low VAF mutations hotspots, presenting a powerful tool for cancer diagnosis and monitoring.

Enzymatic colorimetric method for turn-on determination of l-lactic acid through indicator displacement assay.

l-Lactic acid is a natural α-hydroxy carboxylic acid and is commonly used as an addictive. Quantitation of l-lactic acid is indispensable in food and cosmetic industries. An enzymatic colorimetric method was developed for the determination of l-lactic acid by competitive indicator displacement assay. Boric acid inhibited the colorimetric reaction of l-3,4-dihydroxyphenylalanine (l-DOPA) catalyzed by tyrosinase. l-Lactic acid competitively displaced and released l-DOPA bound with boric acid to serve as substrate, and thus restored the tyrosinase activity. Recovery of color reaction could be spectrophotometrically determined at 475 nm and was proportional to the amount of l-lactic acid. A calibration curve between l-lactic acid concentration and recovery of absorbance were built. The concentration range of the l-lactic acid was 0.25-2.25 mM. The limit of detection (LOD) and the limit of quantification (LOQ) for l-lactic acid was estimated to be 0.05 mM and 0.16 mM, respectively. The method achieved turn-on and visual sensing with good precision, accuracy, specificity, and robustness. The assay method exhibited a promising prospect to determine the content of l-lactic acid in foods and cosmetics.

First derivative synchronous fluorometric method to continuously measure monophenolase activity.

Tyrosinase plays an essential role in melanin biosynthesis and inherently exhibits both monophenolase and diphenolase activity. A first derivative synchronous fluorometric assay was established for directly monitoring monophenolase activity. The zero-crossing point at 322 nm for the first-derivative under synchronous fluorescence with Δλ = 67 nm was utilized to selectively quantify tyrosine in the presence of the reaction product dihydroxyphenylalanine (DOPA). The limit of detection (LOD) for tyrosine was 0.54 µM. The fluorescence intensity of tyrosine was monitored at intervals of 30 s to establish the time course of tyrosine consumption. The LOD for the monophenolase activity was 0.0706 U⋅ mL-1. The Michaelis-Menten e constant and maximum speed were 21.83 µM and 1.12 µM min-1, respectively. Zinc ions competitively inhibited the monophenolase activity, with an IC50 value of 14.36 µM. This assay is easily and rapidly executed and is of great significance for analyzing the kinetics of enzymatic reactions and in fundamental research on monophenolase. This approach has potential applications in the discovery of tyrosinase inhibitors for medicine and cosmetics, as well as in the industrial synthesis of substituted o-diphenol intermediates.

The complete chloroplast genome of Cuscuta australis R. Br. (Convolvulaceae) and its phylogenetic implication.

The genus Cuscuta (Convolvulaceae) is an annual parasitic twining herb. There are about 200 species in this genus, which are widely distributed in tropical and subtropical areas. Cuscuta is mainly parasitic on crops bringing significant losses to the production of agriculture. Furthermore, dried seeds of C. chinensis and C. australis are used as a Chinese traditional herbal medicine. Despite the importance of Cuscuta species, it is difficult to distinguish these plants by the naked eye. Moreover, plastid sequence information available for Cuscuta species is limited. In this study, the complete chloroplast (cp) genome sequence of C. australis was determined using next-generation sequencing. The entire cp genome was determined to be 85,263 bp in length. It contained large single-copy (LSC) and small single-copy (SSC) regions of 50,384 and 6727 bp, respectively, which were separated by a pair of 14,076 bp inverted repeat (IR) regions. The genome contained 98 genes, including 61 protein-coding genes, 29 tRNA genes, and 4 rRNA genes. The overall GC content of the genome is 37.8%. A phylogenetic tree reconstructed by 26 chloroplast genomes reveals that C. australis is most related with Cuscuta pentagona in Convolvulaceae, with bootstrap support values of 100%.

[Characteristics of paddy field nitrogen leakage and runoff in rice-duck farming system].

A field experiment was conducted to study the characteristics of paddy field nitrogen (N) leakage and runoff under rice-duck farming (MRD), conventional farming (MR), and conventional farming with flooding (CK). Comparing with that under MR, the paddy field under MRD had a notable decrease of N (especially NO3- -N) concentration in its leaked liquid; but this concentration was tended to be increased, compared with that under CK. After 7-9 days of fertilization, the NH4+ -N and NO3- -N concentrations in paddy field surface water were higher under MRD than under MR. However, owing to the no draining and the higher band, the paddy field under MRD had a notable reduction of drainage, resulting in a marked decrease of N runoff than that under MR. Comparing with MR, the paddy field under MRD had an addition of nitrogen supply from duck dung, a reduction of N leakage and runoff, a lesser application of chemical nitrogen fertilizer, and more nitrogen uptake by rice plant. Both the reduction of N input and that of N output in rice-duck farming system were nearly equal in quantity.