In this work, boronic ester-mediated dual recognition has been coupled with a CRISPR/Cas12a system; thus, a new method for highly specific and sensitive detection of lipopolysaccharide (LPS) is proposed via the simultaneous recognition of boronic acid and an LPS aptamer (LPSA) as well as signal amplification by CRISPR/Cas12a. Specifically, boronic acid-modified magnetic beads (MB@APBA) and aptamers are employed for the simultaneous dual recognition of LPS, while polymerase isotherm amplification is further utilized to induce LPS cycling and form a double strand, which can activate the CRISPR/Cas12a system so as to amplify the signal. Consequently, a linear detection range can be obtained from 0.05 to 5000 ng/mL, with the lowest detection limit of 44.86 pg/mL. The capturing of MB@APBA on 1, 2- and 1, 3-cis dihydroxyl-containing substances can not only eliminate the interference of other molecules but also enhance the highly specific recognition of LPSA on LPS. Moreover, MB@APBA can be reused by adjusting the pH value of the reaction system. The method can be developed as a universal platform for the analytical detection of other carbohydrates.