Background: Core promoters have a substantial influence on various steps of transcription, including initiation, elongation, termination, polyadenylation, and finally, translation. The characterization of core promoters is crucial for exploring the regulatory code of transcription initiation. However, the current understanding of insect core promoters is focused on those of Diptera (especially Drosophila) species with small genome sizes. Results: Here, we present an analysis of the transcription start sites (TSSs) in the migratory locust, Locusta migratoria, which has a genome size of 6.5 Gb. The genomic differences, including lower precision of transcription initiation and fewer constraints on the distance from transcription factor binding sites or regulatory elements to TSSs, were revealed in locusts compared with Drosophila insects. Furthermore, we found a distinct bimodal log distribution of the distances from the start codons to the core promoters of locust genes. We found stricter constraints on the exon length of mRNA leaders and widespread expression activity of the distant core promoters in locusts compared with fruit flies. We further compared core promoters in seven arthropod species across a broad range of genome sizes to reinforce our results on the emergence of distant core promoters in large-sized genomes. Conclusions: In summary, our results provide novel insights into the effects of genome size expansion on distant transcription initiation.