Construction of eukaryotic expression vector containing the gene of insulin-like growth factor-1

摘要

Objective: to establish a recombinant retroviral vector containing IGF-1 gene and to provide the basis for the application of IGF-1 in treating nervous system disease such as stroke. Methods: the plasmid pcDNA3.1-IGF-1 was cut by EcoR I/Xho I, and subcloned to retroviral vector pLXSN, resulting in the recombinant plasmid pLX-IGF-1. This IGF-1 expression vector was evaluated by enzymatic digestion and sequencing. By the Lipofectamin 2000, pLX-IGF-1 was transferred to packaging cell line PA317. Culture supernatant of these cells was detected for titeration of the recombinant virus. Results: We establish a retroviral vector containing IGF-1 gene successfully, and it was proved by enzyme cutting and sequence analysis. The vector producing cell line PA317/IGF-1 was established. Average titers of the recombinant virus in the culture supernatant are about 6.5x10~5 CFU/ml. Conclusion: A recombinant retroviral containing IGF-1 gene was successfully constructed. Insulin-like growth factor-1 (IGF-1) is a peptide factor, which is of alternative nutrition for neurons. It is of nutrition and protection in vivo for various kinds of neuron and affects the survival, growth, proliferation and differentiation of neurons. The amount of IGF-1 decreases with age. The common injection, for the short time period of IGF-1 in brain and than IGF-1 can not pass by the blood-brain barrier, needs times' of injection which will result in the new damage to neurons in the brain. Gene therapy is a very hopeful solution to resolve this problem. For the low efficiency of transfection of brain tissues with plasmids, a high efficient vector is wanted for therapy of central neuronal system diseases with the implantation of target cells. For consideration of this, we manage to construct a eukaryotic expression vector containing the gene of insulin-like growth factor-1.