One-tube smart genetic testing via coupling isothermal amplification and three-way nucleic acid circuit to glucometers

Abstract

Urgent demand for portable diagnosis has promoted a new sensing strategy that uses personal glucometer (PGM) to detect non-glucose targets. Even though great progresses have been achieved in terms of target range and sensing principle, issues such as low final signal-to-background ratio and hard-to-realize one-tube smart analysis still exist and challenge real-world applications in gene detection. Here we propose a practical solution via coupling isothermal amplification (i.e. LAMP) and three-way amplifiable catalytic hairpin assembly (i.e. CHA) to a PGM. It allows direct transduction from genomic information to commercial portable devices with all of ultra-high sensitivity, specificity and enhanced signal-to-noise ratio. Compared with previous report without signal amplification, the introduction of CHA has successfully improved the signal amplitude by at least 12.5 folds. More importantly, through importing an effective three-way junction based transduction, we also innovatively develop a one-tube logical or multiplex analysis strategy in PGM based detection. Totally four situations of two foodborne bacteria genes, in Cronobacter sakazakii (ompA) and Escherichia coli (malB), could be directly readout using the final PGM signals, with the lowest detection amount down to less than 100 molecular copies (6.6 × 10−18 M). It is believed such a LAMP-CHA-PGM method has been already sensitive, specific, and of great potential for practically portable gene diagnostics.