Construction of Complete Telomere-to-Telomere Genome Assembly of the Rabbit Using Haploid Embryonic Stem Cells

A, Y.; Mao, J.; Hu, S.; Yu, M.; Yang, Y.; Zhang, Y.; Qiu, T.; Quan, L.; Zhao, H.; Zhang, Q.; Yu, K.; Ye, Y.; Yan, H.; Li, N.; Yang, X.; Zhang, R.; Zhai, G.; Wang, M.; Lai, L.

doi:10.1101/2025.03.26.645123

bioRxiv · 10.1101/2025.03.26.645123

Construction of Complete Telomere-to-Telomere Genome Assembly of the Rabbit Using Haploid Embryonic Stem Cells

A, Y.·Mao, J.·Hu, S.·Yu, M.·Yang, Y.·Zhang, Y.·Qiu, T.·Quan, L.·Zhao, H.·Zhang, Q.·Yu, K.·Ye, Y.·Yan, H.·Li, N.·Yang, X.·Zhang, R.·Zhai, G.·Wang, M.·Lai, L.

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Abstract

The rabbit owns commercial importance in meat and fur production, and also has long served as a valuable animal model in biomedical research. Yet, the complete assembly of a high-quality rabbit reference genome has not been established. Here, we present a telomere-to-telomere (T2T) genome assembly of the New Zealand White (NZW) rabbit, the most complete and accurate rabbit genome to date. Using a haploid embryonic stem cells (haESCs) to overcame the challenges of homologous sequence interference and structural variation, we generate a genome with a contig N50 of 137.71 Mb, anchoring all chromosomes with only three gaps, through integrating long-read sequencing (PacBio HiFi, Oxford Nanopore), Hi-C, and Illumina data. Quality assessments reveal near-complete coverage of the BUSCO mammalian gene set, with 99.61% completeness. Notably, this complete assembly allowed for the first comprehensive annotation of the rabbit immunoglobulin loci and major histocompatibility complex region, which were previously unresolved in earlier genome versions. The NZW-T2T assembly based on haESCs not only provides a critical resource for advancing rabbit-based research, but also set a new benchmark for de novo genome assembly for other animals with complex genome.

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A, Y., Mao, J., Hu, S., Yu, M., Yang, Y., Zhang, Y., Qiu, T., Quan, L., Zhao, H., Zhang, Q., Yu, K., Ye, Y., Yan, H., Li, N., Yang, X., Zhang, R., Zhai, G., Wang, M., Lai, L.. 2025-03-28. Construction of Complete Telomere-to-Telomere Genome Assembly of the Rabbit Using Haploid Embryonic Stem Cells. https://doi.org/10.1101/2025.03.26.645123

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