文献类型：期刊文献

英文题名：CRISPR/Cas12a and immuno-RCA based electrochemical biosensor for detecting pathogenic bacteria

作者：Chen, Zhibao[1,2];Ma, Li[1,2,3];Bu, Shengjun[3];Zhang, Wenguang[3];Chen, Jinjun[1];Li, Zhongyi[3];Hao, Zhuo[3];Wan, Jiayu[3]

机构：[1]Guangdong Ocean Univ, Coll Coastal Agr Sci, Zhanjiang 524088, Peoples R China;[2]Guangdong Ocean Univ, Shenzhen Res Inst, Shenzhen 518108, Peoples R China;[3]Chinese Acad Agr Sci, Changchun Vet Res Inst, Changchun 130122, Peoples R China

年份：2021

卷号：901

外文期刊名：JOURNAL OF ELECTROANALYTICAL CHEMISTRY

收录：SCI-EXPANDED(收录号：WOS:000709805200002)、、EI(收录号：20214211023418)、Scopus(收录号：2-s2.0-85116909739)、WOS

语种：英文

外文关键词：CRISPR/Cas12a-based detection; Probe based electrochemical biosensor; E. coli O157:H7; Trans-cleavage; RCA

外文摘要：Rapid, simple, and sensitive detection of food-borne pathogenic bacteria is extremely important for preventing and controlling food-borne diseases. Herein, an electrochemical biosensor based on CRISPR/Cas12a combined with immuno-rolling circle amplification (immuno-RCA) was developed for detecting the pathogenic bacterium, Escherichia coli O157:H7. Based on a sandwich-type immunoassay on magnetic beads, immuno-RCA generated long single-stranded DNA with abundant E. coli O157:H7-specific aptamers and target repeated sequences, which allowed recognition and binding of E. coli O157:H7 and complex formation by CRISPR/ Cas12a and crRNA, respectively. Stimulation of Cas12a produced trans-cleavage activity toward a non-specific methylene blue labeled DNA hairpin probe on the electrode surface. In the presence of E. coli O157:H7, the CRISPR/Cas12a non-specific trans-cleavage activity was triggered, the hairpin DNA on the Au electrode was cleaved, and the peak current was altered. Under optimal conditions, the developed biosensor presented a broad dynamic detection range from 10 to 107 CFU center dot mL(-1), with a detection limit of 10 CFU center dot mL(-1). Moreover, the biosensor did not exhibit any cross-reactivity with other non-target bacteria. These results revealed that the developed biosensor is a simple, sensitivity, and specific platform for E. coli O157:H7 detection.