Transposable elements (TEs) are mobile DNA sequences, colloquially known as ‘jumping genes’ because of their ability to replicate to new genomic locations. Given a vector of transfer (e.g. tick or virus), TEs can jump further: between organisms or species in a process known as horizontal transfer (HT). Here we propose that LINE-1 (L1) and Bovine-B (BovB), the two most abundant TE families in mammals, were initially introduced as foreign DNA via ancient HT events. Using a 503-genome dataset, we identify multiple ancient L1 HT events in eukaryotes and provide evidence that L1s infiltrated the mammalian lineage after the monotreme-therian split. We also extend the BovB paradigm by increasing the number of estimated transfer events compared to previous studies, finding new potential blood-sucking parasite vectors and occurrences in new lineages (e.g. bats, frog). Given that these TEs make up nearly half of the genome sequence in today’s mammals, our results provide the first evidence that HT can have drastic and long-term effects on the new host genomes. This revolutionizes our perception of genome evolution to consider external factors, such as the natural introduction of foreign DNA. With the advancement of genome sequencing technologies and bioinformatics tools, we anticipate our study to be the first of many large-scale phylogenomic analyses exploring the role of HT in genome evolution.Significance statementLINE-1 (L1) elements occupy about half of most mammalian genomes (1), and they are believed to be strictly vertically inherited (2). Mutagenic L1 insertions are thought to account for approximately 1 of every 1000 random, disease-causing insertions in humans (4-7). Our research indicates that the very presence of L1s in humans, and other therian mammals, is due to an ancient transfer event – which has drastic implications for our perception of genome evolution. Using a machina analyses over 503 genomes, we trace the origins of L1 and BovB retrotransposons across the tree of life, and provide evidence of their long-term impact on eukaryotic evolution.