Yang Ying, Yang Jun
Key Laboratory of Bioactive Materials, Ministry of Education, College of Life Sciences, Nankai University, Tianjin 300071, China.
Sheng Wu Yi Xue Gong Cheng Xue Za Zhi. 2009 Dec;26(6):1286-90.
In order to trace the expression of exogenous hepatocyte growth factor (HGF) in eukaryotic cells exactly, a recombinant plasmid that expresses the fusion protein of HGF and red fluorescent protein (RFP) was constructed. The gene encoding HGF without stop code was amplified from pMD19-T-HGF by PCR technique and then cloned in pDsRed-express-N1. The recombinant plasmid (pDsRed-HGF) was identified by restriction endonuclease enzyme analysis and DNA sequence analysis. pDsRed-HGF was transfected into 293T cells. The expression of HGF mRNA, HGF and red fluorescent protein was detected. The results showed that the target gene sequence in pDsRed-HGF was completely in conformity with HGF cDNA in GenBank, and it was expressed highly in 293T cells. In this study, pDsRed-HGF was successfully constructed and expressed in eukaryotic cells.
为了准确追踪外源性肝细胞生长因子（HGF）在真核细胞中的表达情况，构建了一种表达HGF与红色荧光蛋白（RFP）融合蛋白的重组质粒。通过PCR技术从pMD19-T-HGF中扩增出不含终止密码子的HGF编码基因，然后克隆到pDsRed-express-N1中。通过限制性内切酶分析和DNA序列分析对重组质粒（pDsRed-HGF）进行鉴定。将pDsRed-HGF转染到293T细胞中。检测HGF mRNA、HGF和红色荧光蛋白的表达。结果表明，pDsRed-HGF中的靶基因序列与GenBank中的HGF cDNA完全一致，并且在293T细胞中高表达。在本研究中，pDsRed-HGF成功构建并在真核细胞中表达。
