Transposable elements (TEs) comprise a large portion of eukaryotic genomes. Their capacity to jump within and between genomes contributes to genome plasticity and genetic diversity. However, their role in transcriptional regulation has been less studied, primarily because of the notion that TEs represent “junk”. Understanding TE insertion characteristics, and how their introduction and expansion is controlled, represent central questions in evolutionary biology. Trichomonas vaginalis, a haploid protist whose genome is composed of ~60% TEs provides an excellent system to address these questions. The majority of TEs in this microbe are DNA transposons, including a family of ~1,000 Mariner elements and ~3,000 Maverick elements, which use a cut-and paste mechanism to jump. First we undertook a pilot study to determine in silico insertion preference for these two TE families in a reference genome assembly of T. vaginalis, and found that the interruption of gene expression is inversely correlated with distance of TEs from genes. Surface protein gene families, such as the BspA family, were most frequently disrupted by TE insertions, which might be related to the parasite’s plasticity to establish or modulate infection. Next we looked at insertions across the whole genome using “Transposon Display-Seq” and RNA-Seq of 17 geographically and phenotypically distinct T. vaginalis isolates, and identified a total of 1,411 Maverick and 1,385 Mariner insertions. Approximately 1.7% of Maverick and 0.5% of Mariner insertions were fixed in all samples, some of which showed abolished gene expression in all isolates. Our study represents the first genome-wide analysis of the impact of TE insertions on the T. vaginalis transcriptional landscape and shows that TEs play an important role in gene regulation of this parasite.