DOI: 10.5061/dryad.j0zpc86bk
title: Data from: An annotated draft genome of the mountain hare (Lepus timidus)

description: Hares (genus Lepus) provide clear examples of repeated and often massive
 introgressive hybridization and striking local adaptations. Genomic
 studies on this group have so far relied on comparisons to the European
 rabbit (Oryctolagus cuniculus) reference genome. Here, we report the first
 de novo draft reference genome for a hare species, the mountain hare
 (Lepus timidus), and evaluate the efficacy of whole-genome re-sequencing
 analyses using the new reference versus using the rabbit reference genome.
 The genome was assembled using the ALLPATHS-LG protocol with a combination
 of overlapping pair and mate-pair Illumina sequencing (77x coverage). The
 assembly contained 32,294 scaffolds with a total length of 2.7 Gb and a
 scaffold N50 of 3.4 Mb. Re-scaffolding based on the rabbit reference
 reduced the total number of scaffolds to 4,205 with a scaffold N50 of
 194 Mb. A correspondence was found between 22 of these hare scaffolds and
 the rabbit chromosomes, based on gene content and direct alignment. We
 annotated 24,578 protein coding genes by combining ab-initio predictions,
 homology search, and transcriptome data, of which 683 were solely derived
 from hare-specific transcriptome data. The hare reference genome is
 therefore a new resource to discover and investigate hare-specific
 variation. Similar estimates of heterozygosity and inferred demographic
 history profiles were obtained when mapping hare whole-genome
 re-sequencing data to the new hare draft genome or to alternative
 references based on the rabbit genome. Our results validate previous
 reference-based strategies and suggest that the chromosome-scale hare
 draft genome should enable chromosome-wide analyses and genome scans on
 hares.
description: DNA Sampling, Extraction, and Sequencing One
 female mountain hare (<em>Lepus timidus hibernicus</em>)
 specimen (NCBI BioSample ID SAMN12621015) was captured from the wild for
 scientific research purposes by the Irish Coursing Club (ICC) at
 Borris-in-Ossory, County Laois under National Parks & Wildlife
 (NPWS) license no. C 337/2012 issued by the Department of Arts, Heritage
 and the Gaeltacht (dated October 31, 2012). Genomic DNA was extracted from
 kidney, muscle, and ear tissue using the JETquick Tissue DNA Spin Kit
 (GENOMED), with RNAse and proteinase K to remove RNA and protein
 contamination. Genomic libraries of different insert lengths were
 generated following the standard ALLPATHS-LG protocol (Gnerre et al. 2011): one
 Illumina TruSeq DNA library of 180 bp fragments was sequenced with
 overlapping paired-end (OPE) reads, and three Illumina TruSeq DNA
 mate-pair (MP) libraries of 2.5, 4.5, and 8.0 kb insert sizes.
 Whole-genome sequencing was performed at The Genome Analysis Center (TGAC,
 currently Earlham Institute, Norwich, UK)—seven HiSeq2000 lanes (five OPE
 and two 4.5 kb MP)—and CIBIO’s New-Gen sequencing platform—three HiSeq1500
 lanes (2.5 and 8.0 kb MP). Raw sequencing reads were deposited in the
 Sequence Read Archive. Genome Assembly
 De novo assembly was performed using ALLPATHS-LG (Gnerre
 et al. 2011) with default parameters using OPE and mate-pair
 reads. The resulting assembly was evaluated with REAPR v1.0.18
 (Hunt et al. 2013) to break incorrect scaffolds, by
 mapping the paired-end and the 4.5 kb mate-pair reads on the assembled
 genome. Another round of scaffolding was then performed using SSPACE v3.0
 (Boetzer et al. 2011), with a minimum overlap of 32 bp
 and supported by a minimum of 20 reads
 (CIBIO-ISEM_LeTim1.0_Assembled.fasta.gz). Finally, we
 leveraged the existence of the high-quality assembly of the genome of the
 European rabbit (<i>Oryctolagus cuniculus—Ensembl
 OryCun2.0</i>), to improve the contiguity of the assembly using the
 reference-based scaffolder MeDuSa v.1.6 with five iterations
 (Bosi et al. 2015)
 (CIBIO-ISEM_LeTim1.0_re-scaffolded.fasta.gz). This
 re-scaffolding orders and re-orientates scaffolds without affecting
 intra-scaffold sequence. Quality control of the assembly at different
 stages was assessed based on metrics obtained with QUAST v.3.2
 (Mikheenko et al. 2016). The completeness of
 the <i>L. timidus</i> re-scaffolded genome was evaluated using
 BUSCO v.3.0.2 (Simão et al. 2015), based on the
 presence and absence of core single-copy genes (from mammalia_odb9
 database). We then checked consistency of gene content in the larger
 chromosome-like scaffolds and rabbit chromosomes using blastp from NCBI
 BLAST v2.7.1+ (Camacho et al. 2009), considering the
 best hit per gene with similarity above 90% over 500 bp. The 22 rabbit
 chromosomes were aligned against inferred
 corresponding <i>L. timidus</i> re-scaffolded scaffolds using
 D-Genies v. 1.2.0 Mashmap (Cabanettes and Klopp
 2018). Genome Annotation
 Repetitive regions were identified using RepeatModeler v.1.0.11
 (Smit and Hubley 2008) and masked using RepeatMasker
 v.4.0.7 (Smit et al. 2013). The masked genome was used
 as input for gene prediction in MAKER v.3.01.02 (Cantarel et al.
 2008), using ab-initio
 predictions, <i>L. timidus</i> transcriptome data, and rabbit
 protein annotations (<i>O. cuniculus</i>) -
 (CIBIO-ISEM_LeTim1.0.cdna.abinitio.fa.gz and
 CIBIO-ISEM_LeTim1.0.pep.abinitio.fa.gz). Functional inference for genes
 and transcripts was performed using the translated CDS features of each
 coding transcript. Each predicted protein sequence was based on blastp
 searches against the Uniprot-Swissprot database to retrieve gene name and
 function, and InterProscan v5.30-69 (Jones et al. 2014)
 to retrieve Interpro, Pfam v31.0 (Finn et al. 2016), GO
 (Mi et al. 2017), KEGG (Kanehisa et al.
 2016), and Reactome (Fabregat et al. 2018)
 information (annotation files: CIBIO-ISEM_LeTim1.0_re-scaffolded.gff.gz
 and CIBIO-ISEM_LeTim1.0_re-scaffolded.id.map.gz).
description: This assembly and annotation files corresponds to the version
 CIBIO-ISEM_LeTim1.0 reported in the article "An Annotated Draft
 Genome of the Mountain Hare (<em>Lepus
 timidus</em>)"