•A one-step signal amplified lateral flow strip was developed for on-site detection of mercury ion.•Nucleic acid based signal amplification technique was successfully applied in lateral flow strip based methods.•Extraordinary sensitivity was realized for rapid and ultrasensitive detection of mercury ions.•A potential practical tool for food safety detection and environmental monitoring.•A generous signal amplification technique for nucleic acid probe based lateral flow strip.A novel one-step signal amplified lateral flow strip (SA-LFS) is described for direct ultrasensitive and on-site detection of Hg2+ based on the specific recognition system of thymine–Hg2+–thymine using gold nanoparticles (AuNPs) as labeling tags. The signal was also amplified by hybridization induced dual labeling of AuNPs, which increased signal intensity of test line on strips. The presence of both probes on the LFS enabled both sensing and signal amplification to be achieved in a single step. The resulting SA-LFS is capable of rapid and ultrasensitive detection of Hg2+ with LODs of 0.005 ppb and 0.0015 ppb by visual observation and quantitative analysis, respectively. At least 40-fold improvement in sensitivity of the traditional LFS was achieved through this signal amplification process. The successful application of the method to the determination of Hg2+ in water sample is reported. The proposed signal amplification process may also be applicable to other LFS-based methods.