Deep learning enables the use of ultra-high-density array in DNBSEQ.

DNBSEQ employs a patterned array to facilitate massively parallel sequencing of DNA nanoballs (DNBs), leading to a considerable boost in throughput. By employing the ultra-high-density (UHD) array with an increased density of DNB binding sites, the throughput of DNBSEQ can be further expanded. However, the typical imaging system of the DNBSEQ sequencer is unable to resolve adjacent DNBs spaced smaller than the resolution limit, resulting in poor base-calling performance of the UHD array and hindering its practical application. In this study, we propose a deep-learning-based DNB image super-resolution network named DNBSRN to address this problem. DNBSRN has a specifically designed structure for DNB images and employs a histogram-matching-based preprocessing approach. For the eight DNB image datasets generated from the DNBSEQ sequencer using UHD arrays with 360 nm pitch, the base-calling performances are significantly improved after super-resolution reconstruction by DNBSRN and reached a comparable level to those of the regular density array. In terms of reconstruction speed, DNBSRN takes only 7.61 ms for an input image with 500 × 500 pixels, which minimizes its influence on throughput. Furthermore, compared with state-of-the-art super-resolution networks, DNBSRN demonstrates superior performance in terms of both the quality and speed of DNB image reconstruction. DNBSRN successfully addresses the DNB image super-resolution task. Integrating DNBSRN into the image analysis workflow of DNBSEQ will allow for the application of UHD array, hence enabling a considerable improvement in throughput as well as tremendous savings in unit reagent cost.