Construction of eukaryotic plasmid expressing human TGFBI and its influence on human corne

Construction of eukaryotic plasmid expressing human TGFBI and its influence on human corneal

epithelial cells

佚名

【期刊名称】《《国际眼科杂志：英文版》》 【年(卷),期】2012(000)001

【摘要】AIM: To detect the expression of transforming growth factor beta-induced gene (TGFBI) protein in human corneal tissue and overexpress it in the human corneal epithelial cells in order to discuss the function of TGFBI in the pathogenesis of corneal dystrophy. METHODS: Immunohistochemistry(IHC) was used to detect the expression of TGFBI in the human cornea tissue. TGFBI cDNA was obtained by reverse transcription-PCR from human corneal total RNA extracted from cornea transplant donor and cloned into pCMV-N-HA vector. The recombinant pCMV-N-HA-TGFBI plasmid transfected human corneal epithelial cells. Forty-eight hours later, mRNA and proteins were harvested from cells for real-time PCR analysis and western blot assay respectively.RESULTS: IHC indicated TGFBI mainly exist below the human corneal epithelium layer. Transfection of recombinant pCMV-N-HA-TGFBI into human corneal epithelial cells resulted in effective expression of TGFBI, as shown by increased mRNA level detected by real-time PCR as well as increased protein level detected by Western blot. Meanwhile