人酸性成纤维细胞生长因子真核表达载体的构建及鉴定

Construction and identification of eukaryotic expression vector for human acidic fibroblast growth factor

目的构建人酸性成纤维细胞生长因子真核表达载体pEGFP-N1-haFGF。方法 PCR扩增人酸性成纤维细胞生长因子(haFGF)基因,BglⅡ和HindⅢ双酶切后克隆入真核表达载体pEGFP-N1,构建haFGF真核表达载体pEGFP-N1-haFGF,并经限制性酶谱分析和DNA测序对重组表达载体的正确性进行鉴定。结果重组真核基因表达载体pEGFP-N1-haFGF经限制性内切酶BglⅡ和HindⅢ酶切,DNA电泳显示465 bp的haFGF目的片段和4 700 bp的pEGFP-N1载体片段,重组载体经DNA测序显示所克隆的haFGF基因核苷酸顺序与Genbank中记载的haFGF序列一致。结论本实验成功构建人酸性成纤维细胞生长因子真核表达载体,为进一步研究haFGF的功能奠定基础。

Objective To construct and identify eukaryotic expression vector for human acidic fibroblast growth factor（haFGF）.Methods The haFGF gene was amplified by PCR,digested by BglⅡ and HindⅢ,then the fragment was cloned into eukaryotic expression vector pEGFP-N1.The correctness of recombinant plasmid was identified by restriction endonuclease digesting and DNA sequencing.Results The eukaryotic expression vector for haFGF was digested by Bgl Ⅱ and Hind Ⅲ,and the electrophoresis of the digested products showed two fragments：4 700 bp fragment and 465 bp fragment.The result of DNA sequencing analysis was consistent with the sequence published in GeneBank.Conclusion The eukaryotic expression vector for human acidic fibroblast growth factor pEGFP-N1-haFGF has been constructed and it will establish the base for studying its function and activity in future.