Development of highly effective gene knock-in procedure using neural crest-specific gene expression machinery

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To identify the neural crest cell-specific enhancer region as well as transcription factors that bind to the enhancer region, we conducted luciferase assay using several upstream promoter regions on endothelin type-A receptor gene locus. Consequently, we found that about 30 bases as a critical enhancer sequence that consisted of presumable tandem binding sites of two transcription factors. In vivo experiments in transgenic mice exhibited an insufficiency of this enhancer sequence for neural crest cell-specific forced expression of LacZ gene. These results suggest that a possible coordination with other region may be required to proper gene expression in neural crest cells.Adenoviral transduction in fertilized eggs exhibited toxicity for their later normal development, whereas strong expression of LacZ was observed. Therefore, the important issue of viral toxicity remained to be overcome.