Assembly-graph-assisted haplotype-resolved scaffolding of polyploid genomes

Assembling polyploid genomes at the chromosome level remains a major challenge due to the difficulty of accurately distinguishing homologous chromosomes, leaving a vast number of polyploid species without high-quality genome sequences. Current chromatin conformation capture (3C)-based haplotype-resolved scaffolding methods often produce highly incomplete and mis-phased assemblies of complex polyploids, as short sequences lacking sufficient interaction support and collapsed regions with misleading signals are difficult to resolve. Here we present GPhase, a haplotype-resolved scaffolding algorithm that integrates 3C (for example, Hi-C, Pore-C and Omni-C) data with assembly graphs to enhance accuracy, completeness and contiguity. By leveraging graph connectivity derived from long-read overlaps, GPhase effectively overcomes key limitations of 3C. When applied to 15 complex polyploid genomes, GPhase reduced the sequence missing rate (BUSCO-based) by up to tens-fold, decreased the sequence error rate (k-mer-based) by up to an order of magnitude and improved the contiguity (N50) of gap-free sequences by several-fold compared with existing tools.