Objective In our previous study, we had generated various zebrafish mutant lines with tissue-specific GFP expression by Tol2 transposon-mediated insertional mutagenesis.Among these mutants,the Tol2:20141221t line expresses GFP in nervous system, while the position within zebrafish genome where transposon inserted has not yet been identified. The aim of this study was to identify and analyze this genetically modified mutant.Methods The transgenic insertion loci in the genome of Tol2:20141221t line was identified by TAIL-PCR and the spatial and temporal expression profile of the affected gene was examined by in situ hybridization.Homozygous mutant of Tol2:20141221t was generated for exploring related developmental defects. Results Tol2 transposon was inserted into the 8th intron region of sat1.a gene, and induced premature transcription termination.The maternal and zygotic mutants of Tol2:20141221t was generated, while without apparent developmental defects. Conclusions We have generated and identified the zebrafish sat1.a mutant mediated by Tol2 transposon.This gene insertion mutant exhibits no obvious developmental abnormalities, but may serve as a powerful tool to study the development of nervous system.