Rapid Visualized Detection Of Escherichia Coli O157. H7 By Dna Hydrogel Based On Rolling Circle Amplification

Foodborne illnesses caused by pathogenic bacteria are prominent issues in food safety. Fast and sensitive detection of pathogenic bacteria is significantly essential for food safety. In this work, an aptasensor based on aldehyde magnetic beads, rolling circle amplification (RCA) and DNA hydrogel was developed for visualized, simple and rapid detection of Escherichia Coli O157:H7 (E. coli O157:H7). The Mbs@ dsDNA complexes were first prepared. In the presence of target, the aptamer combined with E. coli O157-H7, which released E. coliaptamer-initiator. The supernatant collected by magnetic separation and the released primers were hybridized with circular sequence to form padlock probe. Then, the RCA reaction was initiated by adding T4 DNA ligase, phi29 DNA polymerase and dNTPs, while the RCA could not be triggered in the absence of target. Those two circular sequences were designed to contain partially complementary bases. Thus, when those two circular sequences generated two long single-stranded DNA (ssDNA) products respectively after RCA reaction, two ssDNA products hybridized with each other and formed naked-eye visible DNA hydrogel. The aptasensor was successfully used for visualized detection of E. coli O157:H7 with a detection limit of 4x10(3) CFU/mL, and the detection could be finished within 30 min. The aptasensor had many advantages such as high specificity and specificity, easy operation, efficient amplification, and rapid and visualized readout, showing potential in applications of food safety detection.