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Dual-reference projection defines a callable core and reference-sensitive shell in a Qiantang River Megalobrama draft
Kai Liu1, Wei Zhang2, Jialing Qiao2
1Institute of Fishery Science, Hangzhou Academy of Agricultural Sciences, Hangzhou, 310024, China. kai@bequ.net.
Functional & Integrative Genomics
|July 31, 2026
Summary
This study assesses projecting short-read fish genome data onto chromosome-level references, revealing that reference choice significantly impacts anchored span but not core sequence integrity. Findings guide interpretation of fragmented genome drafts.
Area of Science:
- Genomics
- Bioinformatics
- Aquatic Biology
Background:
- Short-read genome assemblies in non-model fish often lack chromosome-scale resolution.
- Interpreting these assemblies in chromosome-scale coordinates requires robust projection methods.
- The Qiantang River Megalobrama (SJF) sample provides a case study for evaluating projection accuracy.
Purpose of the Study:
- To evaluate the impact of different chromosome-level reference genomes on the projection of short-read fish genome data.
- To characterize genome architecture and assess the quality of short-read substrates.
- To define the reliable limits of interpretation for fragmented genome drafts.
Main Methods:
- Projecting a polished Illumina-derived genome assembly onto two distinct Megalobrama amblycephala chromosome-level references (NCBI and Genome Warehouse).
- Utilizing k-mer spectra for genome size, heterozygosity, and repeat content estimation.
- Employing NextPolish refinement to improve draft assembly quality.
- Assessing Benchmarking Universal Single-Copy Orthologs (BUSCO) completeness and gene-interval partitioning.
Main Results:
- Estimated haploid genome size of 906.7-909.0 Mb with 1.18-1.20% heterozygosity and 34.7% repeat content.
- NextPolish refinement improved consensus and read support while preserving draft-level BUSCO completeness.
- Reference choice significantly affected the anchored span (919.17-919.24 Mb for reference A vs. 878.98-879.06 Mb for reference B).
- High 50-kb concordance (92.47-96.95%) and similar callable fractions (77.24-77.35%) were observed across references.
- A large callable core (724.85-727.35 Mb) and a smaller reference-sensitive shell were identified.
Conclusions:
- Reference genome selection critically influences the anchored span of projected short-read assemblies.
- A substantial callable core sequence exists, supporting reliable local interpretation in fragmented drafts.
- The study delineates a 'reference-sensitive shell' where interpretation is less supported, guiding future genomic analyses in non-model organisms.
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