From Field to Genome: Non-Lethal, On-Site Nanopore-Only Assembly of the Endangered Grenada Frog (Pristimantis euphronides)

Pristimantis euphronides genome project - data and analysis deposit This deposit contains the complete data, scripts, results, and documentation for the genome assembly, annotation, and analysis of *Pristimantis euphronides* (Grenada Frog, specimen GrenadaFrog144), accompanying the manuscript: Kopp K, Lownds S, Harrison B, Kuhl H, Moittie S. From Field to Genome: Non-Lethal, On-Site Nanopore-Only Assembly of the Endangered Grenada Frog (Pristimantis euphronides). BMC Genomics. In submission. Analysis scripts are additionally maintained at GitHub: https://github.com/koppk/Grenada-Frog-144 Summary The Grenada Frog (Pristimantis euphronides) is a critically endangered direct-developing anuran endemic to montane rainforest on Grenada in the Lesser Antilles. We present a 1.75 Gb scaffold-level reference genome assembled from a single wild female sampled non-lethally by toe clip, blood swab, and buccal swab. All wet-laboratory steps, including Oxford Nanopore Technologies (ONT) MinION sequencing, were performed in-country on portable equipment, and downstream analyses ran on laptop-class hardware and a remote server accessed from Grenada. The Flye-Medaka assembly, scaffolded with RagTag against Eleutherodactylus coqui, comprised 13 pseudochromosomes with a scaffold N50 of 117.8 Mb, achieved 91.4% tetrapod BUSCO completeness, and supported 28,071 annotated protein-coding genes. Cross-species synteny analysis against three additional Hyloidea genomes computationally predicted an underlying karyotype of n = 16, consistent with prior cytogenetic counts. Coverage analysis identified candidate Z-linked regions on two scaffolds and a set of unplaced contigs with sequence features consistent with the heterochromatic W chromosome described from C-banding, providing a first-pass framework for sex chromosome analyses in Pristimantis. Scope This deposit contains input data, output files, intermediate results, figures, tables, and per-directory documentation (including tool versions, parameters, and reproducibility notes). Each analysis directory includes a README with full methods, file descriptions, prerequisites, and cross-references to other sections. Analysis scripts are included alongside the data but are also maintained in the companion GitHub repository. Deposit structure The zip archives listed below follow the sequential order of the manuscript sections. Each zip preserves the internal directory structure including a chapter-specific README. All chapter READMEs are additionally collected in READMEs.zip for quick reference, and a standalone overview README is provided as README_GrenadaFrog.md. README_GrenadaFrog.md Overall README for the repository READMEs.zip Collection of all section README files Reads_FAST.zip FAST (real-time) basecalled Nanopore reads Reads_HAC.zip HAC (high-accuracy) basecalled Nanopore reads Read_quality_analysis.zip Read quality metrics (NanoComp, SeqKit) and Phred score computation Genome_size_and_complexity_estimation.zip Jellyfish k-mer profiling and GenomeScope 2.0 genome size estimation Genome_assembly.zip De novo assemblies: Workflow 1 (Flye/Medaka) and Workflow 2 (Wtdbg) Assembly_quality_assessment.zip Inspector, Merqury, BUSCO (HANNO), per-contig QC, and inter-workflow concordance Taxonomic_classification_of_reads.zip Kraken2 contamination screening against Hyloidea and PlusPF databases Reference_genome_selection_via_k-mer-based_distance_estimation.zip Mash distance estimation and heatmap visualization Reference_genome_selection_via_phylogenetic_proximity.zip Phylogenetic reference selection using Portik et al. (2023) time tree Reference-guided_scaffolding_of_genome_assemblies.zip RagTag and Ragout scaffolding against E. coqui, WindowMasker, contig placement statistics Evaluation_of_reference-guided_scaffolding.zip D-Genies pairwise synteny, contig allegiance, karyotype reconstruction Read_mapping_and_genome_coverage_analysis.zip Read mapping, mosdepth coverage computation, anomaly detection Alignment_with_previously_published_Pristimantis_euphronides_ and_Eleutherodactylus_johnstonei_sequences.zip Minimap2 and BLASTn alignment against published P. euphronides and E. johnstonei sequences Annotation_scaffolded_genome_assembly.zip HANNO gene annotation (pre- and post-scaffolding), NCBI/Zenodo reformatting, GFF3 and FASTA outputs Identification_of_candidate_sex-chromosome-linked_regions.zip Z- and W-candidate regions, gametolog discovery, banding analysis, DM-domain search Phylogenetic_analysis_using_nuclear_gene_orthologs.zip 18-gene extraction, MAFFT alignment, IQ-TREE supermatrix and single-gene phylogenies Divergence_times_and_biogeographic_context.zip Divergence time estimation, AmphibiaWeb geographic enrichment, type-locality distances Haplotype_analysis.zip HapDup haplotype phasing, independent scaffolding of both haplotypes, inter-haplotype concordance Survey_of_anuran_genome_assemblies_in_NCBI_Genome.zip Survey of 214 anuran genome assemblies classified by technology, tissue, and taxonomy Figures.zip Main manuscript and supplementary figures (PNG, PDF, TIFF) with caption files Tables.zip Main manuscript and supplementary tables (ODT, PDF) with caption files Additional_files.zip Additional files 1–12 as submitted to BMC Genomics (supplementary methods, results, tables, figures, and survey) Each analysis directory contains a scripts, inputs and outputs subfolder, and a chapter-specific README. Data availability Raw sequencing reads: ENA / NCBI BioProject: PRJEB89028 Assembly: ENA / NCBI (aPriEup1.0): GCA_965278355.2 Analysis scripts: GitHub: koppk/Grenada-Frog-144