Droplet-based nucleic acid amplification in a temperature gradient

摘要

A method of amplifying a nucleic acid, the method comprising cycling an amplification-ready droplet through a temperature gradient to locations within the temperature gradient suitable for effecting steps in an amplification reaction.

主权项

1. A method of amplifying a nucleic acid, the method comprising:
(a) providing an amplification-ready droplet loaded operations gap of a droplet actuator, wherein the droplet actuator comprises:
(i) two substrates separated to provide the droplet operations gap therebetween;(ii) electrodes associated with the substrates and arranged for transporting droplets in the droplet operations gap by electrowetting, the electrodes including an electrode path comprising electrodes arranged to transport droplets along the electrode path; and(iii) an oil-based filler fluid in the droplet operations gap at least partially surrounding the amplification-ready droplet; (b) applying heat to the droplet actuator to establish a temperature gradient of gradually varying temperature in the droplet operations gap, wherein the temperature gradient is established using a heating element arranged to heat filler fluid at a locus of the electrode path; and (c) cycling the amplification-ready droplet by using the electrodes to transport the amplification-ready droplet through the temperature gradient to locations within the temperature gradient having temperatures suitable for effecting steps in an amplification reaction, wherein the droplet is shuttled along the electrode path to different locations within the temperature gradient, and improved amplification patterns based on the results of varying the locations are identified.