Massively Parallel Selection of NanoCluster Beacons

Abstract

NanoCluster Beacons (NCBs) are multicolor silver nanocluster probes whose fluorescence can be activated or tuned by a proximal DNA strand called the activator. While a single-nucleotide difference in a pair of activators can lead to drastically different activation outcomes, termed the polar opposite twins (POTs), it is difficult to discover new POT-NCBs using the conventional low-throughput characterization approaches. Here we report a high-throughput selection method that takes advantage of repurposed next-generation-sequencing (NGS) chips to screen the activation fluorescence of ~40,000 activator sequences. We find the nucleobases at positions 7-12 of the 18-nucleotide-long activator are critical to creating bright NCBs and positions 4-6 and 2-4 are hotspots to generate yellow and red POTs, respectively. Based on these findings, we propose a "zipper bag model" that explains how these hotspots lead to the creation of distinct silver cluster chromophores and contribute to the difference in chromophore chemical yields. Combining high-throughput screening with machine learning algorithms, we establish a pipeline to rationally design bright and multicolor NCBs.

Beyond nature's base pairs: machine learning-enabled design of DNA-stabilized silver nanoclusters

“…Copp, et al , developed the high-throughput experimental approach described in Section 4, 43 and the Yeh group has developed a platform for rapid screening of “light-up” Ag N –DNA probes called NanoCluster Beacons (NCBs) 80 based on Illumina MiSeq chips. 81…”

“…9). 108 Using a next-generation sequencing chip platform to screen nearly 40 000 activator strands on a fluorescence microscope equipped with TRITC and Cy5 filter cubes, 81 they identified thousands of new activator sequences that yield bright NanoCluster Beacon fluorescence. This dataset was then used to train logistic regression classifiers to distinguish between DNA activator sequences that light-up brightly fluorescent “yellow-orange” and red NanoCluster Beacons and DNA activator sequences associated with low fluorescence (“dark”).…”