High-Quality Genomic DNA Extraction and Sequencing

In order to extract high-quality gDNA, we used the QIAGEN Blood & Cell Culture DNA Kit. We then selected the high molecular weight gDNA (targeting 10–50 kb) using a Blue Pippin system (Sage Science, Beverly, MA, USA) and further processed the Nanopore sequencing library with the Ligation sequencing 1D kit (SQK-LSK108, ONT, UK) according to the manufacturer’s instructions. We sequenced the resulting library through the GridION X5 sequencer (ONT, UK) at the Genome Center of Nextomics (Wuhan, China). Base calling was further carried out on fast5 files using the ONT Albacore software v0.8.4, and low-quality reads (mean_qscore <7) and adapter sequences were filtered. Sequencing libraries were also prepared with gDNA using Illumina Genomic DNA Sample Preparation Kit and sequenced on an Illumina HiSeq X Ten system in paired-end mode (2 × 150 bp). Adapter sequences and low-quality reads were removed using NGS QC Toolkit v 2.3.3.¹⁷ (link) The obtained clean data were used for error correction and k-mer analysis. The Hi-C library was prepared from 3 g of freshly ground young leaves, using liquid nitrogen with a mortar and pestle. The chromatin extraction, digestion, DNA ligation, purification, and fragmentation were all performed as previously described.¹⁸