ObjectiveA mammalian Delta-Notch signaling component,Notch1, has been suggested for its expression during the normal sperm development although its conditional deletion caused no apparent abnormalities. Since we established our original transgenic mouse system that enabled labeling of past and ongoingNotch1signaling at a cellular level, we tried to validate that observation in vivo. Our transgenic mouse system used Cre/loxP system to induce tandem dsRed expression upon Notch1 signaling.ResultsTo our surprise, we were unable to observe tandem dsRed expression in the seminiferous tubules where the sperms developed. In addition, tandem dsRed expression was lacking in the somatic cells of the next generation in our transgenic mouse system, suggesting that sperms received noNotch1signaling during their development. To validate this result, we conducted re-analysis of four single-cell RNA-seq datasets from mouse and human testes and showed thatNotch1expression was little in the sperm cell lineage. Collectively, our results posed a question into the involvement ofNotch1in the normal sperm development although this observation may help the interpretation of the previous result thatNotch1conditional deletion caused no apparent abnormalities in murine spermatogenesis.Supplementary InformationThe online version contains supplementary material available at 10.1186/s13104-023-06378-z.