High-throughput single-cell DNA methylation and chromatin accessibility co-profiling with SpliCOOL-seq

DNA methylation and chromatin accessibility are fundamental epigenetic mechanisms that orchestrate gene expression programs, define cellular states, and drive developmental trajectories. scCOOL-seq has enabled simultaneously measuring the two modalities in the same single cells, but in quite a low throughput manner. We present single-cell split-pool ligation-based multi-omics sequencing technology (SpliCOOL-seq), which improves the throughput to thousands of cells by combining split-pool ligation based single-cell indexing after in situ tagmentation with universal Tn5 transposase and scCOOL-seq. SpliCOOL-seq achieved higher sensitivity than previous high throughput single-cell DNA methylation sequencing methods and can clearly distinguish different lung cancer cells based on both genetic and multiple epigenetic modalities. We show that the two DNMT inhibitors, 5-Azacitidine and Decitabine, both cause large scale demethylation but in distinct patterns. Applied to the primary lung tumor, SpliCOOL-seq clearly captured subclones within the tumor lesion and revealed candidate genes related to tumorigenesis. Furthermore, we presented the first report on the heterogeneity of scDNAm age acceleration among tumor subclones as predicted from a single-cell perspective. In conclusion, SpliCOOL-seq achieves parallel profiling of whole genome DNA methylation and chromatin accessibility in the same individual cells in a high-throughput manner and is hopefully used to illustrate regulatory interactions under different cell states.