Crispr-cas 12a combination to alleviate the false-positive in loop-mediated isothermal amplification-based diagnosis of Neisseria meningitidis [post]

Ngo Tat Trung, Le Huu Phuc Son, Trinh Xuan Hien, Dao Thanh Quyen, Mai Hong Bang, Le Huu Song
2020 unpublished
Loop mediated Isothermal amplicafication (LAMP) was recently suggested as a diagnostic tool for the identification of Neisseria meningitides. Howevere, this isothermal amplification is challenged by the fact its amplification leads to risks of obtaining false-positive results. Whereas, with abilities to accurately recognize specific sequence, the CRISPR/Cas12a can forms complexes with cognate RNA sensors and cleave pathogen's DNA targets complimerntary to its cognate RNA and acquires collateral
more » ... acquires collateral activity to unbiasedly cut nearby off-target fragments. Therefore, if relevant fluorescent-quencher-nucleic probes are present in the reaction, the non-specific cleavage of probes releases fluorescences and establish diagnostic read-outs. In this study, we demonstrate a proof-of-concept that in relevant biochemical conditions, CRISPR/Cas12a and LAMP can work synchronously to identify genetics materials of Nesseria menitigistis at the level of 0.00004% in less than 2 h. Additionally, our clinical data also showed that the combinatory of CRISPR/Cas12a help to alleviate false positive result therefore enhance the specificity gained by the LAMP assays.
doi:10.21203/rs.3.rs-108208/v1 fatcat:hztzlqxlhjgn3cr6jzq7tdblou