Liping Xia,Juxin Yin,Jianjian Zhuang,Weihong Yin,Zheyu Zou,Ying Mu
Rapid and sensitive pathogen detection methods are critical for disease diagnosis and treatment. RPA-CRISPR/Cas12 systems have displayed remarkable potential in pathogen detection. A self-priming digital PCR chip is a powerful and attractive tool for nucleic detection. However, the application of the RPA-CRISPR/Cas12 system to the self-priming chip still has great challenges due to the problems of protein adsorption and two-step detection mode of RPA-CRISPR/Cas12. In this study, an adsorption-free self-priming digital chip was developed and a direct digital dual-crRNAs (3D) assay was established based on the chip for ultrasensitive detection of pathogens. This 3D assay combined the advantages of rapid amplification of RPA, specific cleavage of Cas12a, accurate quantification of digital PCR, and point-of-care testing (POCT) of microfluidics, enabling accurate and reliable digital absolute quantification of Salmonella in POCT. Our method can provide a good linear relationship of Salmonella detection in the range from 2.58 × 101 to 2.58 × 104 cells/mL with a limit of detection ∼0.2 cells/mL within 30 min in a digital chip by targeting the invA gene of Salmonella. Moreover, the assay could directly detect Salmonella in milk without nucleic acid extraction. Therefore, the 3D assay has the significant potential to provide accurate and rapid pathogen detection in POCT. This study provides a powerful nucleic detection platform and facilitates the application of CRISPR/Cas-assisted detection and microfluidic chips.
Jiajie Xu,Zhiyu Liu,Tenghui Jin,Xusheng Yang,Huihuang Chen,J. Paul Chen
Fenton reaction has been widely used for efficient treatment of organic wastewater. However, its applications are limited by such key factors as pH < 3. In this study, we developed, tested, and optimized an alginate/C 3 N 4 porphyrin bead (C 3 N 4 por-SA) as a recyclable photocatalyst in a photocatalysis-self-Fenton process to overcome these limitations. Porphyrin-modified C 3 N 4 (C 3 N 4 por) was used as the H 2 O 2 donator, while Fe(III) nodes served as the Fenton reagent . The as-prepared floating alginate/C 3 N 4 por bead utilized the light source as a driving force for the catalysis. Under visible light irradiation for 6 h, the model pollutant atrazine was degraded by 70.96 % by the optimized photocatalyst (named as C 3 N 4 por-SA-Fe1Ca5), demonstrating better photocatalytic performance than alginate/C 3 N 4 beads. This improvement was attributed to the higher H 2 O 2 yield from C 3 N 4 por. The alginate/C 3 N 4 por bead showed better photocatalytic activity even after several consecutive cycles and could easily be recovered for reuse. Furthermore, Fe(III)/Ca(II) bimetallic alginate bead exhibited better photocatalytic activity and a higher content of •OH radicals than the Ca(II) monometallic alginate beads, due to the ability of Fe(III) nodes to serve as a Fenton reagent. The influences of light sources, and commonly existing matters (namely SO 4 2− , Cl − , CO 3 2− , NO 3 − , and humic acid) were investigated. Moreover, the alginate/C 3 N 4 por bead demonstrated good photocatalytic performance in a simulated natural environment without the addition of extra H 2 O 2 , with an atrazine removal percentage of up to 96.3 % after 3-h irradiation. These findings indicated the great potential of alginate/C 3 N 4 por bead in practical applications.
危险品化学品经营许可证（带存储）