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SunUp and Sunset genomes revealed impact of particle bombardment mediated transformation and domestication history in papaya

Nature Genetics volume 54, pages 715–724 (2022)Cite this article

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Abstract

Transgenic papaya is widely publicized for controlling papaya ringspot virus. However, the impact of particle bombardment on the genome remains unknown. The transgenic SunUp and its progenitor Sunset genomes were assembled into 351.5 and 350.3 Mb in nine chromosomes, respectively. We identified a 1.64 Mb insertion containing three transgenic insertions in SunUp chromosome 5, consisting of 52 nuclear-plastid, 21 nuclear-mitochondrial and 1 nuclear genomic fragments. A 591.9 kb fragment in chromosome 5 was translocated into the 1.64 Mb insertion. We assembled a gapless 9.8 Mb hermaphrodite-specific region of the Yh chromosome and its 6.0 Mb X counterpart. Resequencing 86 genomes revealed three distinct groups, validating their geographic origin and breeding history. We identified 147 selective sweeps and defined the essential role of zeta-carotene desaturase in carotenoid accumulation during domestication. Our findings elucidated the impact of particle bombardment and improved our understanding of sex chromosomes and domestication to expedite papaya improvement.

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Fig. 1: Distribution of genomic features along the SunUp and Sunset genomes.
Fig. 2: Integration site of transgenic insertions in SunUp genome.
Fig. 3: The distribution of NUPTs and NUMTs in SunUp and Sunset genomes.
Fig. 4: Comparison of the gapless HSY and its counterpart X.
Fig. 5: Population structure of cultivated and wild papaya.
Fig. 6: Signatures of artificial selection and evidence of domestication in cultivated papaya.

Data availability

Nanopore and PacBio whole-genome sequencing data, Hi-C, Illumina data and RNA-seq data have been deposited in the NCBI Sequence Read Archive (SRA) database as Bioproject PRJNA727683. The SunUp and Sunset genome assemblies were archived in the NCBI Genome database under the accession number JAIUCH000000000 for SunUp and JAIUCG000000000 for Sunset genome. The SunUp and Sunset genome assemblies and gene annotations have been also deposited in the Genome Warehouse (GWH) database in BIG data Center (https://ngdc.cncb.ac.cn/gwh/) under accession number GWHBFSC00000000 for SunUp genome and GWHBFSD00000000 for Sunset genome. VCF file that contains all clean SNPs was uploaded to the Mendeley database (https://data.mendeley.com/datasets/m5phbmw43c/1). The papaya sex-specific small RNA sequence data used can be obtained from NCBI’s Gene Expression Omnibus (GEO) under accession number GSE54097. Source data are provided with this paper.

Code availability

The Python scripts ‘filter_blast_result.sh’, ‘convert_bed_for_junction.py’, ‘extract_result.py’ and ‘plot_data_V8.py’ for Sunset and SunUp-specific nuclear organelle DNA junction sites identification were available at GitHub (https://github.com/sc-zhang/CUFscripts/tree/v1.0). These codes were also archived on Zenodo with the https://doi.org/10.5281/zenodo.6342427.

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Acknowledgements

This work was supported by US National Science Foundation (NSF) Plant Genome Research Program Award (DBI-1546890 to R.M.), National Natural Science Foundation of China grants (31701889 to J.Y.) and Natural Science Foundation of Fujian Province grants (2018J01601 to J.Y. and 2018J01604 to Xingtan Zhang). The Science and Technology Innovation Fund of Fujian Agriculture and Forestry University (CXZX2020091A to J.Y).

Author information

Author notes

  1. These authors contributed equally: Jingjing Yue, Robert VanBuren, Juan Liu, Jingping Fang.

Authors and Affiliations

  1. Center for Genomics and Biotechnology, Fujian Provincial Key Laboratory of Haixia Applied Plant Systems Biology, Key Laboratory of Genetics, Breeding and Multiple Utilization of Crops, Ministry of Education, Fujian Agriculture and Forestry University, Fuzhou, China

    Jingjing Yue, Juan Liu, Jingping Fang, Xingtan Zhang, Zhenyang Liao, Xiuming Xu, Shuai Chen, Shengchen Zhang, Xiaokai Ma, Yaying Ma, Hongying Yu, Jing Lin, Ban Deng, Fang Deng, Xiaobing Zhao, Hansong Yan, Mahpara Fatima, Zhicong Lin, Haibao Tang & Jisen Zhang

  2. Department of Plant Biology, University of Illinois at Urbana-Champaign, Urbana, IL, USA

    Robert VanBuren, Ching Man Wai, Dessireé Zerpa-Catanho, Xiaodan Zhang & Ray Ming

  3. Department of Horticulture, Michigan State University, East Lansing, MI, USA

    Robert VanBuren

  4. Key Laboratory of Plant Germplasm Enhancement and Specialty Agriculture, Wuhan Botanical Garden, Chinese Academy of Sciences, Wuhan, China

    Juan Liu & Mei Yang

  5. College of Life Science, Fujian Normal University, Fuzhou, China

    Jingping Fang

  6. Key Laboratory of the Ministry of Education for Coastal and Wetland Ecosystems, College of the Environment and Ecology, Xiamen University, Xiamen, China

    Xiuming Xu

  7. Fruit Research Institute, Fujian Academy of Agricultural Sciences, Fuzhou, China

    Ping Zhou

  8. Institute of Oceanography, Minjiang University, Fuzhou, China

    Yongji Huang

  9. Department of Tropical Plant and Soil Sciences, University of Hawaii at Manoa, Honolulu, HI, USA

    Nancy J. Chen & Robert E. Paull

  10. Estación Experimental Agrícola Fabio Baudrit Moreno, Universidad de Costa Rica, Alajuela, Costa Rica

    Eric Mora-Newcomer & Patricia Quesada-Rojas

  11. Estación Experimental Los Diamantes, Instituto de Innovación y Transferencia de Tecnología Agropecuaria, Guápiles, Costa Rica

    Antonio Bogantes

  12. CIGRAS, Universidad de Costa Rica, San Pedro, Costa Rica

    Víctor M. Jiménez

  13. Hawaii Agriculture Research Center, Kunia, HI, USA

    Ming-Li Wang

  14. Texas A&M AgriLife Research, Texas A&M University System, Dallas, TX, USA

    Qingyi Yu

Authors
  1. Jingjing Yue
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  2. Robert VanBuren
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  3. Juan Liu
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  9. Shuai Chen
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Contributions

R.M. conceived this genome project and coordinated the research activities. R.M., J.Y., R.V. and Q.Y. designed the experiments. J.Y., R.V., Juan Liu and J.F. carried out the experiments. Xingtan Zhang., X.X., H. Yan and Z. Liao assembled SunUp and Sunset genomes. J.Y., Z. Lin., B.D., Xiaobing Zhao., Xiaodan Zhang and F.D. processed genome sequencing and resequencing data. J.Y., Juan Liu., Q.Y., M.Y., Xingtan Zhang., Y.H., Z. Liao and J.F. analyzed transgenic insertions. J.Y., Jing Lin., X.X., S.Z., P.Z., Y.M., H. Yu., J.F., M.F., J.Z., H.T. and M.-L.W. analyzed SunUp and Sunset genome and RNA-seq data. R.V., C.M.W., S.C., N.J.C., R.E.P., E.M.-N., P.Q.-R., A.B., V.M.J., X.M. and D.Z.-C. maintained plant materials and analyzed resequenced genomes. J.Y., R.V. Juan Liu., J.F. and R.M. wrote the manuscript.

Corresponding author

Correspondence to Ray Ming.

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Competing interests

The authors declare no competing interests.

Peer review

Peer review information

Nature Genetics thanks Aureliano Bombarely, Jordi Garcia-Mas and the other, anonymous, reviewer(s) for their contribution to the peer review of this work.

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Extended data

Extended Data Fig. 1 Genome-wide analysis of chromatin interactions in papaya SunUp and Sunset genomes.

Hi-C interactions heatmap diagram at 500Kb resolutions for the entire nine chromosomes and HSY region were showed from top left to bottom right in order (1–9, indicate Chr1-Chr9, HSY indicates hermaphrodite-specific Yh chromosome region). The color bars beside heat maps indicate strong interactions in red and weak interactions in yellow.

Extended Data Fig. 2 PCR validation of transgenic insertion sites in Sunset.

a, The schematic diagram for the 1.64 Mb insertion in SunUp. Primers were designed to amplify the insertion loci P3, and sequences of both sides were defined as P1 and P2 in SunUp and Sunset. b, The insertion loci P3 can be amplified in SunUp but not in Sunset. PCR products for band P3 were confirmed by sanger-sequencing, and the result was consistent with assembly sequence. The experiment was repeated three times and the same results were gotten. c-d, The dot plot for both border sequences in SunUp and Sunset which were marked as Sunset P1/P2 and SunUp P1/P2.

Extended Data Fig. 3 Validation of the nuclear sequences located on 1.64 Mb insertion by Nanopore reads in SunUp.

The nuclear DNA fragment located in Chr5: 37.29-37.31 Mb of SunUp was mapped to the Sunset genome with the corresponding region in Chr5: 37.04-37.06 Mb of Sunset. The assembly of the nuclear DNA fragment was verified by Nanopore reads. SunUp Nanopore reads were aligned against the SunUp reference genome using the BWA software, and the result was visualized by Jbrowse.

Extended Data Fig. 4 Pipeline of identification of SunUp-specific genomic integration of nuclear organelle DNA fragments.

a, Quality control of raw sequenced data. b, Search for SunUp nuclear organelle junction sites by BLASTN. c, Alignment between Sunset reads and SunUp reference genome. Unmapped reads were removed in further analysis. d, Nuclear organelle junction sites shared by SunUp and Sunset. A junction site was supposed to be shared by SunUp and Sunset genomes when there were mapped to reads and spanning its position in the SunUp reference genome. e, Extraction of shared reliable junction sites. f, Junction sites specific in SunUp.

Extended Data Fig. 5 Comparison of the 591.9 Kb deletion with SunUp, chloroplast and mitochondria genomic sequences by using BLASTN.

The 591.9 Kb deletion (Chr5: 22.0-22.6 Mb) in Sunset was aligned to SunUp Chr5, chloroplast and mitochondrial sequences. The plot indicates that the 591.9 Kb deletion has the corresponding region in SunUp Chr5: 36.4-38.1 Mb, chloroplast and mitochondrial sequences.

Extended Data Fig. 6 Model of integration of exogenous DNA into the papaya genome.

By comparison between SunUp and its host Sunset genome, our results supported a precisely regulated model that essential events may be involved in integration of exogenous DNA into the plant genome via particle bombardment. Three transgenic insertions were integrated into a single site locus. Particle bombardment mediated transformation brought foreign DNA fragments into the NUPT-rich region, accompanied by the integration of chloroplast and mitochondrial genome fragments into nuclear genome and the translocation and rearrangement of the original NUPTs and NUMTs. Organellar DNA fragments were transferred to nuclear genome by the driving force. This force may be penetration of cells with DNA-coated metal particles that elicits a wound response. This wound response would activate DNA repair and degradation enzymes on the introduced DNA, making transgenic sequences and NUPTs and NUMTs rearrangement.

Extended Data Fig. 7 PCR validation of transgenic insertions.

Three transgenic insertions, 9789 bp functional insertion including CP, 1533 bp tetA insertion and 290 bp nptII insertion were amplified in Sunset and SunUp. Three insertions can be amplified in SunUp but not in Sunset by using the 5 primer pairs listed in Supplementary table 66. The experiment was repeated three times and the same results were obtained. PCR products were confirmed by Sanger-sequencing, and the result was used to correct the assembly sequence.

Extended Data Fig. 8 Validation of the insertion sites by PacBio and Nanopore reads in SunUp.

Aligning the PacBio reads and Nanopore reads of SunUp and Sunset against the transgenic function fragment using BWA software, and the results were shown by Jbrowse. Part of SunUp reads were displayed on the region. No Sunset reads were spanned the functional insertion.

Extended Data Fig. 9 The position of PCR-amplified regions on the insertion.

We designed 65 pairs of primers which were randomly distributed on the 1.64 Mb insertion of SunUp (SunUp Chr5: 36.42-38.06 Mb) to amplify insertion sequences in SunUp and Sunset. The information of primers were shown in Supplementary table 66. Thick lines are shown on the insertion region, with blue and red lines indicating amplified bands. All blue and red bands can be amplified in SunUp genome, and blue lines indicate SunUp-specific fragments that can be only amplified in SunUp not in Sunset.

Supplementary information

Supplementary Information

Supplementary Tables 1–11, 17, 22–29, 33–35, 37–39, 41, 42, 44, 47, 50, 51, 57, 60, 62–65, 67 and 68, and Figs. 1–11.

Reporting Summary

Supplementary Tables

Supplementary Tables 12–16, 18–21, 30–32, 36–40, 43, 45, 46, 48, 49, 52–56, 58, 59, 61 and 66.

Source data

Source Data Fig. 1

Unprocessed gels.

Source Data Fig. 2

Unprocessed gels.

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Yue, J., VanBuren, R., Liu, J. et al. SunUp and Sunset genomes revealed impact of particle bombardment mediated transformation and domestication history in papaya. Nat Genet 54, 715–724 (2022). https://doi.org/10.1038/s41588-022-01068-1

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