Construction and validation of Tg (Crystal-pro-gsdf- 2A-egfp)plasmid in zebrafish
GUO An-ning, GUAN Gui-jun, WANG Ying, SUN Kai-qing, CHEN Liang-biao
Key Laboratory of Exploration and Utilization of Aquatic Genetic Resources, Ministry of Education, International Research Centre for Marine Bioscience, Ministry of Science and Technology, Shanghai Ocean University, Shanghai 201306, China
Abstract: Total RNA extracted from testis of zebrafishDaniorerio was reversely transcribed to cDNA as a template for amplification of gonadal soma-derived factor gene (gsdf) by RT-PCR, and a reporter vector to over-expressgsdf-2A-egfp fusion protein was constructed under the mouse gamma-crystalin promoter in order to investigate the potential role ofgsdf gene playing in the mechanism of testicular differentiation. This reporter(Tg:Crystal-pro-gsdf-2A-egfp)was microinjected into one cell stage of zebrafish embryos and enabled a green fluorescence protein gene (gfp) andgsdf expression in lens of the fish from embryos to adults, with both transgenic lines of zebrafish (F1). Among nineteen adults withgfp signals in eyes found under a fluorescence microscope, specifically expressed green fluorescent protein of F2 generation was finally obtained by hybridization of zebrafish expressing green fluorescent protein with wild-type (WT) non-transgenic zebrafish. The findings indicate thatgsdf -2A-egfp can be inheridated and stable passed into the next generation through germline transmission in zebrafish. The establishment of Tg(Crystal-pro-gsdf-2A-egfp) transgenic line in zebrafish and provided a new method for research and observation of molecular regulation mechanism ofgsdf gene in the development of zebrafish testis.
2018, Vol. 33 
