Yi Jilin, Tian Geng
Department of General Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030.
J Huazhong Univ Sci Technolog Med Sci. 2003;23(4):392-5. doi: 10.1007/BF02829426.
To clone the murine alpha-fetoprotein (AFP) gene, construct the eukaryotic expression vector of AFP and express in CHO cells, total RNA were extracted from Hepa 1-6 cells, and then the murine alpha-fetoprotein gene was amplified by RT-PCR and cloned into the eukaryotic expression vector pcDNA3.1. The recombinant of vector was identified by restriction enzyme analysis and sequencing. After transient transfection of CHO cells with the vector, Western blotting was used to detect the expression of AFP. It is concluded that the 1.8 kb murine alpha-fetoprotein gene was successfully cloned and its eukaryotic expression vector was successfully constructed.
为克隆小鼠甲胎蛋白（AFP）基因，构建AFP的真核表达载体并在CHO细胞中表达，从Hepa 1-6细胞中提取总RNA，然后通过RT-PCR扩增小鼠甲胎蛋白基因并克隆到真核表达载体pcDNA3.1中。通过限制性酶切分析和测序鉴定载体的重组体。用该载体对CHO细胞进行瞬时转染后，用蛋白质免疫印迹法检测AFP的表达。得出结论：成功克隆了1.8 kb的小鼠甲胎蛋白基因并成功构建了其真核表达载体。
