Biological system

Biology

Computer science

Digital polymerase chain reaction

Gene

Biophysics

microRNA

Biochemistry

Thermodynamics

Isothermal process

Computational biology

Nanotechnology

Loop-mediated isothermal amplification

Polymerase chain reaction

Chemistry

Materials science

Physics

Science Advances

MicroRNAs, a class of transcripts involved in the regulation of gene expression, are emerging as promising disease-specific biomarkers accessible from tissues or bodily fluids. However, their accurate quantification from biological samples remains challenging. We report a sensitive and quantitative microRNA detection method using an isothermal amplification chemistry adapted to a droplet digital readout. Building on molecular programming concepts, we design a DNA circuit that converts, thresholds, amplifies, and reports the presence of a specific microRNA, down to the femtomolar concentration. Using a leak absorption mechanism, we were able to suppress nonspecific amplification, classically encountered in other exponential amplification reactions. As a result, we demonstrate that this isothermal amplification scheme is adapted to digital counting of microRNAs: By partitioning the reaction mixture into water-in-oil droplets, resulting in single microRNA encapsulation and amplification, the method provides abso...

Isothermal digital detection of microRNAs using background-free molecular circuit