Genome assemblies were performed for eight wild-type strains of Drosophila melanogaster and Drosophila simulans from Oxford Nanopore long read sequencing (please refer to Mohamed et al. Cells 2020 (doi:10.3390/cells9081776)). Assemblies were deposited in the European Nucleotide Archive (ENA) at EMBL-EBI under accession number PRJEB50024 (https://www.ebi.ac.uk/ena/browser/view/PRJEB50024). Transposable Element annotations: we used RepeatMasker 4.1.0 (http://repeatmasker.org/) -species Drosophila, followed by OneCodeToFindThemAll (Bailly-Bechet et al. 2014) with default parameters. Gene annotations: For D. melanogaster, we retrieved dmel-all-exon-r6.16.fasta, dmel-all-intron-r6.16.fasta, dmel-all-five_prime_UTR-r6.16.fasta, dmel-all-five_prime_UTR-r6.16.fasta, and dmel-all-gene-r6.16.fasta, which are available at ftp.flybase.net/genomes/Drosophila_melanogaster/dmel_r6.16_FB2017_03/fasta/. For D. simulans, we retrieved dsim-all-exon-r2.02.fasta, dsim-all-intron-r2.02.fasta, dsim-all-five_prime_UTR-r2.02.fasta, dsim-all-three_prime_UTR-r2.02.fasta, and dsim-all-gene-r2.02.fasta, which are available at ftp.flybase.net/genomes/Drosophila_simulans/dsim_r2.02_FB2017_04/fasta/. These different sequence sets were aligned to our assemblies using BLAT (Kent 2002) and pslReps from the BLAT suite, in order to keep the best hit (-singleHit). We filtered out genes with 0 hit and genes with multiple hits. For each gene, we also checked that the positions of the hits for each of the internal regions (i.e. exon, intron, 5’ UTR, and 3’ UTR) were included in the positions of the complete gene hit. We excluded regions that did not fulfill these criteria. We also excluded hits that were not on the same chromosome as the corresponding gene on the reference genome.