Abstract
Chromosomes are key structural and functional components of heredity. Reconstruction of ancestral cell karyotypes (ACKs) and evolutionary trajectories of chromosomes (CETs) can reveal how structural and functional changes in chromosomes have occurred during evolution. The whole-genome duplication integrated analysis toolkit implements a telomere-centric model on the basis of the comparative analysis of gene collinearity within and between plant genomes to reconstruct the ACKs and CETs of many angiosperm plants whose genomes have been complicated by repeated polyploidization and subsequent repatterning. Here we summarize the steps for using whole-genome duplication integrated analysis to infer the gene collinearity within a genome or between genomes and to infer the repeated polyploidization specific to a plant or common to multiple plants or plant families. In addition, we provide an example with three grass genomes. We also describe procedures to infer ancestral chromosomes at key evolutionary nodes, to reconstruct CETs from the deep past to extant plants and to generate event-related hierarchical alignment of multiple genomes, which is realized by deciphering collinear genes produced by different polyploidization or speciation events. The Protocol guides users to infer ACKs and CETs in a plant taxon and between selected plants from different taxa, which is crucial to understand important sources of genetic innovation including chromosome evolution, genome complexity and origination and evolution of duplicated genes. This Protocol requires minimal bioinformatics knowledge, for example, retrieving data from public databases and running Python programs. Completing the protocol with the example data takes around 8 h.
Key points
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Whole-genome duplication integrated analysis implements a telomere-centric model of chromosome evolution, whereby telomeres are assumed to have a central role in chromosome structural rearrangements during plant evolution. The tool infers gene collinearity within and between plant genomes and uses it to obtain ancestral cell karyotypes and reconstruct evolutionary trajectories of chromosomes.
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Unlike related tools that rely on gene synteny, whole-genome duplication integrated analysis infers collinear genes that are related to specific polyploidization and speciation events.
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Data availability
Genome sequences and annotations used in this protocol can be retrieved from the NCBI genome (https://www.ncbi.nlm.nih.gov/datasets/genome). The data for this protocol are wrapped as a zip file named ‘WGDI_protocol.zip’, which is available at http://47.92.161.27:8000/biaoge/backend/download.html. Source data are provided with this paper.
Code availability
WGDI is available via GitHub at https://github.com/SunPengChuan/wgdi-example. Programs used in this protocol and example datasets are included within the ‘WGDI_protocol.zip’ file, which is available at http://47.92.161.27:8000/biaoge/backend/download.html.
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Acknowledgements
This work was supported by research grants from the National Natural Science Foundation of China to X.W. (grant no. 32070669) and to Z.W. (grant no. 32000405), the Talented Scientist program from the Tangshan Municipal Bureau Human Resources and Social Security to X.W. (grant no. 16013601) and the Tangshan Science and Technology Program Project (grant no. 21130228) to Z.W.
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X.W. conceived and led the research and prepared the manuscript. B.S., W.L., Y.J., L.Y., T.W., M.Y., J.W., F.H., H.Q., H.Z., Z.W. and Y.J. analyzed the data and prepared the figures; B.S. and X.W. provided the description of WGDI; F.H. and J.W. analyzed the data related to partial monocots and eudicots, respectively, and W.L., Y.J., T.W. and Y.L. improved the programs inferring ACKs and CETs to ensure user-friendly output.
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Nature Protocols thanks Kai Ye and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.
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Key references
Wang, J. et al. Mol. Biol. Evol. 35, 16–26 (2018): https://doi.org/10.1093/molbev/msx242
Sun, P. et al. Mol. Plant 15, 1841–1851 (2022): https://doi.org/10.1016/j.molp.2022.10.018
Wang, S. et al. Genome Biol. 22, 304 (2021): https://doi.org/10.1186/s13059-021-02522-9
Supplementary information
Supplementary Information
Supplementary Figs. 1–4 and text.
Supplementary Data 1
Original data sets for readers to repeat the analysis described in the manuscript and the Supplementary text.
Supplementary Data 2
A demo for the protocol to run WGDI to infer ancestral chromosomes and evolutionary trajectories to form the chromosomes in extant plants.
Supplementary Data 3
Source data for Supplementary Figs. 1–3.
Source data
Source data of Figs. 3c–g and 6
Statistical source data.
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Wang, X., Song, B., Liu, W. et al. Comparative genomics approach to infer ancestral cell karyotypes and reconstruct the evolutionary trajectories of plant chromosomes. Nat Protoc (2025). https://doi.org/10.1038/s41596-025-01173-5
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DOI: https://doi.org/10.1038/s41596-025-01173-5
