Picoliter Well Array Chip-Based Digital Recombinase Polymerase Amplification for Absolute Quantification of Nucleic Acids

Li, Zhao; Liu, Yong; Wei, Qingquan; Liu, Yuanjie; Liu, Wenwen; Zhang, Xuelian; Yu, Yude

We developed a simple and effective scraping liquid blade to partition a sample into as many as 27,000 independent reaction microwells. We scraped liquid with a certain strength to make the soft silica gel contact closely with the chip surface. Thus, after dispensing the reagent effectively, there was lesser residual liquid on the surface for reducing the opportunity of cross- contamination [73] . To produce a large number of monodispersed droplets, the scraping time (~5 s) is much shorter than that of other microdroplet techniques, in which droplets are commonly produced sequentially by T-junctions or flow [37] [38] [39] [40] [41] . Current digital droplet systems take considerable time for droplet production, which leads to amplification in the bulk solution. Thus, they cannot be used to perform dRPA. In contrast, the platform presented here can be applied to many DNA amplification techniques, such as RPA, PCR, RCA, and LAMP. This sample loading style does not require pumping equipment or precise microvalve control [36] .