Sensitive, rapid and low-cost nucleic acid detection is critical for controlling infectious pathogens.Here, we develop a ready-to-use and multimodal detection based on a rebuilding-free, ultrasensitive and selective strategy named dual hairpin ligation-induced isothermal amplification pro (DHLApro). Taking influenza A, influenza B, MERS-CoV, SARS-CoV-2 as model targets, we demonstrate DHLApro provides � zM level ultra-sensitivity, being equaling to 0.45 copy/μL in original sample. By simply changing the recognition module, a set of DHLApro components can be applied to a new target without performance loss. Moreover, DHLApro innovatively allows flexible logic/multiplex assay using one set of primer, for example, the "N pathogens-in-1" OR gate screening and accurate multichannel multiplex assay. Compared with traditional methods, the cost of this logic/multiplex assay has been largely reduced and the cross-interference between the multiple primer sets is also avoided.

Sensitive, rapid and low‐cost nucleic acid detection is critical for controlling infectious pathogens. Here, we develop a ready‐to‐use and multimodal detection based on a rebuilding‐free, ultrasensitive and selective strategy named dual hairpin ligation‐induced isothermal amplification pro (DHLApro). Taking influenza A, influenza B, MERS‐CoV, SARS‐CoV‐2 as model targets, we demonstrate DHLApro provides ≈zM level ultra‐sensitivity, being equaling to 0.45 copy/μL in original sample. By simply changing the recognition module, a set of DHLApro components can be applied to a new target without performance loss. Moreover, DHLApro innovatively allows flexible logic/multiplex assay using one set of primer, for example, the “N pathogens‐in‐1” OR gate screening and accurate multi‐channel multiplex assay. Compared with traditional methods, the cost of this logic/multiplex assay has been largely reduced and the cross‐interference between the multiple primer sets is also avoided.