关键词: Hath1基因, gfp基因, 真核表达, 荧光显微镜

Abstract:

Objective To construct the eukaryotic expression vector of Hath1 gene and to transfect neuroblastoma cells (SH-SY5Y),and to observe its expression in SH-SY5Y cells.Methods DNA was extracted from human whole blood，the Hath1 gene was cloned and the green fluorescent protein(GFP) gfp gene was amplified at the same time; the primers GFP-R and Hath1-F was used to amplify fusion gene gfp-Hath1,and gfp-hath1 was cloned into pMD18-T and digested with XhoⅠ and EcoRⅠ. The recombinant expression plasmid pCDNA3.1 (+) :: gfp-Hath1 was constructed and   transfected into SH-SY5Y cells.Results The gfp-Hath1 fusion gene with 2 023 bp was successfully amplified.The eukaryotic expression plasmid pCDNA3.1 (+) :: gfp-Hath1 was constructed successfully after identified with double enzyme digestion.The GFP was found in SH-SY5Y cells  under immunofluorescence  microscope.Conclusion The Hath1 gene is expressed in SH-SY5Y cells successfully, which lays  a foundation for transfection cochlea experiment and exploring new methods to treat deafness.

Key words: Hath1 gene, gfp gene, eukaryotic expression, fluorescence microscope