Time‐Efficient and Informatic‐Skill‐Light Gap‐Filling for Telomere‐to‐Telomere Genome Assembly

ABSTRACT Despite remarkable advances in sequencing technologies and automated genome assembly algorithms, manual gap‐filling remains indispensable for achieving telomere‐to‐telomere (T2T) genome assemblies, a process that can take weeks or even months. Additionally, these tasks require advanced bioinformatics expertise, thereby excluding many biologists from direct participation in T2T genome projects. This severely restricts the ability to construct T2T genomes for larger populations and a wider range of species. To overcome these challenges, we developed GapSuite, an integrated auxiliary software toolbox that includes two complementary tools, Gap‐Aid and Gap‐Graph, which facilitate gap‐filling through sequence‐extension‐based and assembly‐graph‐based strategies, respectively. The two tools empower users with limited computational expertise to efficiently perform gap closure on personal computers with just mouse clicks, resulting in a fully assembled genome. GapSuite incorporates several technical innovations to achieve key functions and improve both time and space efficiency. Their effectiveness was validated using Arabidopsis thaliana , rice and human genomes as well as simulated diploid and polyploid genomes. As case studies, we used the tools to construct, to the best of our knowledge, the first T2T genome of rice 9311, a model variety of indica rice, and to fill part of the remaining gaps in a recently published gapless poplar genome.