目的：探讨成纤维细胞生长因子4（ FGFR4）重组质粒的构建方法，并检测FGFR4融合蛋白的表达。方法从HepG2细胞中提取总RNA，采用RT-PCR方法扩增FGFR4的全长编码序列，双酶切后与pcDNA3．1／Myc-HisA载体连接，构建pcDNA3．1／Myc-HisA-FGFR4重组质粒。重组质粒经双酶切和测序鉴定后瞬时转染人胚肾HEK293细胞，采用Western blot法检测FGFR4融合蛋白的表达。结果 FGFR4编码序列被成功克隆至pcDNA 3．1／Myc-HisA质粒中，成功构建了pcDNA3．1／Myc-HisA-FGFR4载体。转染HEK293细胞后检测到FGFR4融合蛋白的稳定表达，分子量约为89 kD。结论成功构建了FGFR4全长编码基因重组质粒，并检测到转染后FGFR4蛋白在HEK293细胞中的表达。%Objective To investigate the construction method of the recombinant plasmid of fibroblast growth factor re-ceptor 4(FGFR4) and to identify the expression of its fusion protein.Methods Total RNA was extracted from HepG2 cells.We obtained the full length of human FGFR4 coding sequence by RT-PCR, digested it by restriction enzyme and joined it with pcDNA3.1/Myc-HisA to construct the pcDNA3.1/Myc-HisA-FGFR4 recombinant plasmid.After the recom-binant plasmid was identified by enzyme digestion and sequencing, the plasmid was transfected into HEK293 cells.The ex-pression of fusion protein in HEK293 cells was detected by Western blotting.Results The coding sequence of human FG-FR4 was successfully cloned into pcDNA3.1/Myc-HisA.The pcDNA3.1/Myc-HisA-FGFR4 vector was successfully con-structed.After transfecting HEK293 cells, the stable expression of FGFR4 fusion protein with a molecular weight of 89 kD was detected.Conclusion The recombinant plasmid of FGFR4 full-length coding gene was successfully constructed and the FGFR4 protein was expressed in HEK293 cells.
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