Near complete assembly of Drosophila melanogaster Canton S strain genome

The fruit fly Drosophila melanogaster is a pivotal model organism, yet its reference genome (ISO-1 strain) retains unresolved gaps in complex regions. Here, we present a near-complete telomere-to-telomere genome assembly (Dm.nT2T) of the Canton S strain (male), generated using an integrative approach combining PacBio HiFi, Oxford Nanopore ultra-long reads, and Hi-C data. This assembly spans 161.63 Mb, closes 93.28% of gaps in the current reference genome, and improves contiguity (contig N50 of 21.93 Mb). We identify centromeric and telomeric regions, yielding a more complete genome map. Comparative analysis with the ISO-1 reference identifies 7989 structural variants, including large insertions/deletions enriched in telomeric and centromeric regions. Notably, we detect SINE transposable elements, and identify 92 genes via multi-strategy annotation. Functional validation of two genes shows that Chr3L.2449, expressed in olfactory tissues, is critical for olfactory sensitivity. Dm.nT2T provides a high-quality resource for exploring genomic complexity, strain diversity, and gene function in Drosophila.