{
  "abstract": "De novo transcriptome assembly is the standard approach for constructing a reference transcriptome in non-model plants that lack a high-quality genome, yet short-read assemblies struggle to resolve full-length isoforms. Long-read Iso-Seq (PacBio) captures full-length transcripts directly, but its use as a primary reference and the choice of downstream assembly pipeline remains poorly benchmarked. Here, we construct genome-free Iso-Seq reference transcriptomes for two organs, flower and leaf, of the non-model species Moricandia arvensis (L.) DC. (Brassicaceae), and systematically compare pipeline strategies combining Iso-Seq clustering, CD-HIT redundancy reduction, and Cogent graph-based reconstruction, benchmarked by BUSCO completeness, RSEM short-read mapping, and TransDecoder ORF completeness. We find that the optimal pipeline is organ specific. For flower, CD-HIT pre-filtering followed by Cogent reconstruction produced a high-quality reference (95.3% BUSCO complete). For the leaf, the same Cogent step was detrimental, reducing BUSCO completeness from 90.1% to 78.0% by incorrectly merging distinct genes; therefore, CD-HIT at 95% identity without reconstruction was retained. We trace this divergence to organ-specific input-data characteristics: leaf transcripts show extreme full-length-read expression skew and predominantly single-isoform gene support, depriving Cogent's graph algorithm of the multi-isoform evidence it requires. We find that the concentration of full-length reads among the most highly expressed transcripts predicts pipeline suitability before reconstruction, with per-transcript read depth acting as a necessary but non-discriminating floor. Because the leaf reference lacked gene-level structure, we further recovered gene-isoform grouping using expression-aware read-clustering (Corset), which preserved completeness while restoring the paralog structure expected of a paleopolyploid genome and outperformed sequence-only clustering. Our results provide a robust, genome-free framework for constructing full-length reference transcriptomes in non-model plant species and demonstrate that pipeline choice must be evaluated per organ rather than assuming one size fits all.",
  "abstract_shown": true,
  "affiliations": [
    "Universidad de Granada"
  ],
  "article_type": "New results",
  "arxiv": null,
  "author_total": 8,
  "authors": [
    {
      "name": "S. Sharma"
    },
    {
      "name": "M. Hackenberg"
    },
    {
      "name": "L. Navarro"
    },
    {
      "name": "E. Narbona"
    },
    {
      "name": "A. Gonzalez-Megias"
    },
    {
      "name": "C. Armas"
    },
    {
      "name": "J. M. Gomez"
    },
    {
      "affs": [
        0
      ],
      "name": "F. Perfectti"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/0xned3v3w8bng.bib",
  "cited_by": 0,
  "doi": "10.64898/2026.09.08.750093",
  "fulltext_url": "https://www.biorxiv.org/content/10.64898/2026.09.08.750093v1.full",
  "id": "0xned3v3w8bng",
  "issns": [],
  "landing_url": "https://www.biorxiv.org/content/10.64898/2026.09.08.750093v1",
  "language": null,
  "licence": "CC-BY",
  "licence_label": "CC BY",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "biorxiv",
      "bioinformatics"
    ],
    [
      "openalex",
      "1312|Genomics and Phylogenetic Studies"
    ]
  ],
  "openalex": "W7212195106",
  "pdf_url": "https://www.biorxiv.org/content/10.64898/2026.09.08.750093v1.full.pdf",
  "pmcid": null,
  "pmid": null,
  "published": "2026-09-08",
  "publisher": null,
  "ris": "https://research.draconitic.com/paper/0xned3v3w8bng.ris",
  "sources": [
    {
      "first_seen": "2026-10-30T12:20:42Z",
      "native_id": "10.64898/2026.09.08.750093",
      "source": "biorxiv",
      "url": "https://www.biorxiv.org/content/10.64898/2026.09.08.750093v1"
    }
  ],
  "subjects": [
    {
      "field": "biology",
      "origin": "biorxiv:bioinformatics",
      "subfield": "bioinformatics"
    }
  ],
  "title": "Benchmarking long-read RNA sequencing for de novo transcriptome assembly in non-model plant species: insights from Moricandia arvensis",
  "title_text": "Benchmarking long-read RNA sequencing for de novo transcriptome assembly in non-model plant species: insights from Moricandia arvensis",
  "type": "preprint",
  "updated_at": "2026-10-30T12:20:42Z",
  "url": "https://research.draconitic.com/paper/0xned3v3w8bng",
  "venue": "bioRxiv",
  "versions": []
}