Construction of eukaryotic expression vector with granulocyte-macrophage colony-stimulating factor gene
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Objective: To construct the eukaryotic expression vector that express human granulocyte-macrophage colony-stimulating factor (hGM-CSF) gene for making highly express in mammalian cells. Methods: Extract totally RNA from the induced human fetal lung (HFL) cell line. HGM-CSF cDNA was obtained by reverse transcription-polymerase chain reaction (RT-PCR), and then directionally subcloned into the HindIII and EcoRI site on the pcDNA3.1 plasmid, which was controlled by the CMV promoter, to form the recombinant expressing vector pcDNA3.1-GM-CSF. Results: The PCR amplification was identified and the sequence was analyzed, the results showed that hGM-CSF was properly inserted into the vector and the sequence was correct.
Key wordsHuman granulocyte-macrophage colony-stimulating factor (hGM-CSF) Reverse transcription and polymerse chain reaction (RT-PCR) Eukaryotic expression
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- Yao Jun, Gang RB, Zhang Q, et al. Cloning of human GM-CSF cDNA and expression in E.coli. Acta Biochemica et Biophysica Sinica 1996;28: 265.Google Scholar
- Wu Nai-hu, eds. Principle of Genetic Engineering 2nd ed. 1998; 107.Google Scholar
- Lin Wangming, Yang LF, Huang SZ, et al. PCR Operating and Application Guide. 1st Beijing: People Health Press, 1995.Google Scholar
- Sambrook J, Fritsch EF, Maniatis T. Molecular Cloning, eds. 2nd ed. Cold Spring Harbor Laboratory Press, 1989Google Scholar