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Near telomere-to-telomere genome of the model plant Physcomitrium patens

Nature Plants volume 10, pages 327–343 (2024)Cite this article

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Abstract

The model plant Physcomitrium patens has played a pivotal role in enhancing our comprehension of plant evolution and development. However, the current genome harbours numerous regions that remain unfinished and erroneous. To address these issues, we generated an assembly using Oxford Nanopore reads and Hi-C mapping. The assembly incorporates telomeric and centromeric regions, thereby establishing it as a near telomere-to-telomere genome except a region in chromosome 1 that is not fully assembled due to its highly repetitive nature. This near telomere-to-telomere genome resolves the chromosome number at 26 and provides a gap-free genome assembly as well as updated gene models to aid future studies using this model organism.

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Fig. 1: T2T assembly strategy for the P. patens V6 genome.
Fig. 2: Chromosomal features of the V6 genome.
Fig. 3: Structural conflicts between two distinct versions of particular chromosomes.
Fig. 4: Characterization of centromeres.
Fig. 5: 3D genome attributes of the protonema and gametophore.

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Data availability

The genome assembly and de novo annotations have been deposited in Figshare at https://doi.org/10.6084/m9.figshare.22975925.v2. The Illumina reads (genomic sequencing and Hi-C) and Nanopore reads generated in this study were deposited into the NCBI SRA with the BioProject ID PRJNA742485. The V5 genome assembly was submitted to NCBI with the WGS accession ABEU00000000 under BioProject ID PRJNA13064. The updated gene model lookup table and merged annotation are available for download from https://peatmoss.plantcode.cup.uni-freiburg.de/ppatens_db/downloads.php. The ChIP-seq data generated in this study are available through NGDC (https://ngdc.cncb.ac.cn) with accession PRJCA016808. The V6 genome and annotation data have been submitted to Phytozome (https://phytozome-next.jgi.doe.gov/) and will be made available in their upcoming release.

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Acknowledgements

We thank C. Chen from Huazhong Agricultural University and C. Yu from the Agricultural Genomics Institute at Shenzhen for their advice on chromosome analysis. We also thank H. Chen at Tsinghua University for providing assistance with P. patens genetics. This work was supported by grants from the National Key Research and Development Program of China (no. 2019YFA0906200 to J. Yan), the National Natural Science Foundation of China (nos. 31725002, 32150025 and 32030004 to J.D.), the Bureau of International Cooperation, Chinese Academy of Sciences (no. 172644KYSB20180022 to J.D.), the Shenzhen Science and Technology Program (no. KQTD20180413181837372 to J.D.), the Science Technology and Innovation Commission of Shenzhen Municipality of China (no. ZDSYS20200811142605017 to J. Yan) and the Shenzhen Outstanding Talents Training Fund to J.D. J. Yan acknowledges funding from the Innovation Program of the Chinese Academy of Agricultural Sciences and the Elite Young Scientists Program of CAAS. The gene annotation was carried out in the framework of MAdLand (http://madland.science, DFG priority program 2237). S.A.R. is grateful for funding from the DFG (RE 1697/15–1, 20–1).

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  1. These authors contributed equally: Guiqi Bi, Shijun Zhao, Jiawei Yao, Huan Wang.

Authors and Affiliations

  1. Shenzhen Branch, Guangdong Laboratory of Lingnan Modern Agriculture, Key Laboratory of Synthetic Biology, Ministry of Agriculture and Rural Affairs, Agricultural Genomics Institute at Shenzhen, Chinese Academy of Agricultural Sciences, Shenzhen, China

    Guiqi Bi, Jiawei Yao, Huan Wang, Jianbin Yan & Junbiao Dai

  2. CAS Key Laboratory of Quantitative Engineering Biology, Guangdong Provincial Key Laboratory of Synthetic Genomics and Shenzhen Key Laboratory of Synthetic Genomics, Shenzhen Institute of Synthetic Biology, Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences, Shenzhen, China

    Shijun Zhao, Yingxin Ma & Junbiao Dai

  3. University of Chinese Academy of Sciences, Beijing, China

    Shijun Zhao, Yuanyuan Sun, Xueren Hou & Junbiao Dai

  4. Guangdong Province Key Laboratory of Microbial Signals and Disease Control, Integrative Microbiology Research Centre, South China Agricultural University, Guangzhou, China

    Huan Wang

  5. College of Life Sciences and Oceanography, Shenzhen University, Shenzhen, China

    Mengkai Zhao

  6. State Key Laboratory of Plant Genomics, Institute of Genetics and Developmental Biology, Innovative Academy of Seed Design, Chinese Academy of Sciences, Beijing, China

    Yuanyuan Sun, Xueren Hou & Yuling Jiao

  7. Department of Algal Development and Evolution, Max Planck Institute for Biology Tübingen, Tübingen, Germany

    Fabian B. Haas

  8. Faculty of Chemistry and Pharmacy, University of Freiburg, Freiburg, Germany

    Deepti Varshney & Stefan A. Rensing

  9. Department of Cell and Developmental Biology, University of California, San Diego, La Jolla, CA, USA

    Michael Prigge

  10. School of Life Sciences, Center for Quantitative Biology, and Peking-Tsinghua Center for Life Sciences, Peking University, Beijing, China

    Yuling Jiao

Authors
  1. Guiqi Bi
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  2. Shijun Zhao
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  3. Jiawei Yao
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  4. Huan Wang
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  5. Mengkai Zhao
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  6. Yuanyuan Sun
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  7. Xueren Hou
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  8. Fabian B. Haas
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  9. Deepti Varshney
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  10. Michael Prigge
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  11. Stefan A. Rensing
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  12. Yuling Jiao
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  13. Yingxin Ma
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  14. Jianbin Yan
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  15. Junbiao Dai
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Contributions

J. Yan, Y.M. and J.D. conceived the study. J. Yan and J.D. managed the major scientific objectives. G.B. designed the T2T genome assembly, evaluation and data analysis. J. Yao generated the plant materials. H.W. helped with the assembly and annotation. S.Z., M.Z., Y.S. and X.H. collected the sequenced samples. J. Yao, Y.J., Y.M. and J.D. designed and performed ChIP. F.B.H., D.V., M.P. and S.A.R. combined and quality-checked the gene annotations. J. Yan and G.B. led the article preparation, together with J. Yao, H.W. and J.D. All authors read and approved the final article.

Corresponding authors

Correspondence to Jianbin Yan or Junbiao Dai.

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The authors declare no competing interests.

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Nature Plants thanks the anonymous reviewers for their contribution to the peer review of this work.

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Extended data

Extended Data Fig. 1 The process of graph-based gap-filling for the remaining 16 gaps.

a, Fifteen chromosome karyotype maps exhibit the remaining 16 gaps, which are marked with numerical values. The position of the centromere is the region where the chromosome constriction is situated. b, Upstream and downstream sequences of breakpoints (or gaps) that are aligned with the corresponding graphs. The Hi-C heatmaps accurately reveal gaps (indicated by the intersection of green lines) at a resolution of 1 kb. The upstream sequences (blue band) and downstream sequences (green band) of each gap are aligned with the graphs. On the corresponding pathway, the precise locations of 14 intervals where gaps occur on the graph are highlighted (blue and green bands). The gray region between the two bands represents the sequence that requires filling. Overlapping between the two bands suggests sequence redundancy, necessitating trimming and merging. In the case of gap number 11, its existence is attributed to tandem repeats, demanding estimation of the corresponding copy number before initiating gap repair. c, Two gaps occur in the complex region. The upstream and downstream intervals of two gaps are labeled using four distinct colors. The area is characterized by brief repetitive sequences, and to determine the precise paths for gap-filling, the process of mapping nanopore reads onto this region is utilized. Long reads that are capable of traversing the repetitive structure are then extracted to facilitate path building.

Extended Data Fig. 2 Genome assembly validation achieved by analyzing sequencing coverage and depth in relation to the 26 chromosomes in P. patens.

The coverage (0-100%) and depth information of Illumina and ONT sequencing reads on 26 chromosomes are illustrated in the left and right images, respectively. The statistical analysis was performed using a nonoverlapping window of 50 kbp. Except for the repetitive region adjacent to the centromere of Chr01, which was deliberately omitted from the secondary mapping findings, the ONT reads demonstrated comprehensive coverage of all other chromosomal regions. Furthermore, the sequencing depth of the multicopy rRNA region was markedly elevated, exceeding the typical chromosome sequencing depth of 66x, which further supports the notion of the presence of several copies of rRNA.

Extended Data Fig. 3 Taxonomy distribution obtained by analyzing the assembled unmapped short reads against the NR database using MEGAN6.

The tree represents the taxonomic classification of the matched sequences at the class level, with node size indicating the number of matched sequences. The word cloud displays the sequence matching results at the phylum level, with larger words indicating a greater number of matched sequences.

Extended Data Fig. 4 A fundamental overview of the main content presented in this work.

a, Distribution of 17-23 K-mer frequencies in the P. patens genome. b, A radar chart was utilized to show the quality disparity between the V6 genome and its antecedent, V3. The evaluation was based on six distinct indicators, and the findings were scrutinized to identify any discrepancies in quality between the two versions. c, A concise diagram illustrating the process of V6 assembly. d, Results of SyRI analysis showing genome sequence collinearity and structural variants. To ensure the utmost precision in capturing the genuine discrepancies between the two genome versions, the V3 sequence was fragmented into contigs (where N bases were interrupted). Then, using RaGOO software, 26 pseudochromosomes were created to align with V6.

Extended Data Fig. 5 The neighbor-joining cladogram tree of five P. patens accessions built by SNPs derived from Haas et al.31.

The genome sequencing material used in this study is denoted on the tree by an arrow. Bootstrap values under 100 replicates are shown on nodes.

Extended Data Fig. 6 Assembly accuracy validation for Chr25 in V6 by read mapping.

The above depiction aims to compare the level of collinearity displayed by Chr25 in the V3 and V6 versions. The top section of the diagram portrays the amalgamation of two pseudochromosomes in V3, with their boundaries demarcated by a solid black line. The position of the breakpoint in V6 is indicated by a dashed black line. The middle segment of the diagram illustrates the mapping results of nanopore reads (above 10 kbp). The bottom section of the illustration offers a more comprehensive view of the 5 kbp interval encompassing the breakpoint for detailed scrutiny.

Extended Data Fig. 7 Whole genome-wide Hi-C heatmap.

Hi-C interactions among 26 chromosomes at a 500 kbp resolution.

Supplementary information

Supplementary Information

Supplementary Figs. 1–30 and Note.

Reporting Summary

Supplementary Data 1

Supplementary Tables 1–23.

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Bi, G., Zhao, S., Yao, J. et al. Near telomere-to-telomere genome of the model plant Physcomitrium patens. Nat. Plants 10, 327–343 (2024). https://doi.org/10.1038/s41477-023-01614-7

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