Nanopore Ultra-long Sequencing and Adaptive Sampling Spur Plant Complete Telomere-to-Telomere Genome Assembly

The pursuit of complete telomere-to-telomere (T2T) genome assembly in plants, challenged by genomic complexity, has been advanced by Oxford Nanopore Technologies (ONT), which offers ultra-long, real-time sequencing. Despite its promise, sequencing length and gap filling remain significant challenges. This study optimized DNA extraction and library preparation, achieving DNA lengths exceeding 485 Kb, average N50 read lengths of 80.57 Kb, with reaching up to 440 Kb, and maximum reads of 5.83 Mb. Importantly, it demonstrated that combining ultra-long sequencing and adaptive sampling can effectively fill gaps during assembly, evidenced by successfully achieving the Arabidopsis genome remaining gaps and an unknown telomeric region in watermelon. Our methodologies improve the feasibility of complete T2T genomic assemblies across plants, enhancing genome-based research in diverse fields.