Agarose droplet microfluidics for highly parallel and efficient single molecule emulsion PCR

Abstract

An agarose droplet method was developed for highly parallel and efficient single molecule emulsion PCR. The method capitalizes on the unique thermoresponsive sol–gel switching property of agarose for highly efficient DNA amplification and amplicon trapping. Uniform agarose solution droplets generated via a microfluidic chip serve as robust and inert nanolitre PCR reactors for single copy DNA molecule amplification. After PCR, agarose droplets are gelated to form agarose beads, trapping all amplicons in each reactor to maintain the monoclonality of each droplet. This method does not require cocapsulation of primer labeled microbeads, allows high throughput generation of uniform droplets and enables high PCR efficiency, making it a promising platform for many single copy genetic studies.