Construction of pcDNA3.1(+)GDNF vector and its expression in eukaryotic cells

Objective To construct pcDNA3.1(+)GDNF recombinant eukaryotic expression plasmid and to investigate its expression in eukaryotic cells.Methods The coding sequence of GDNF was amplified from rat astrocytes by reverse transcription PCR (RT PCR) and was cloned into pcDNA3.1(+) eukaryotic expression vector.The recombinant pcDNA3.1(+)GDNF plasmid was then transfected into eukaryotic cells mediated by Fu Gene 6.Analysis by restriction enzyme digestion and DNA sequencing were carried out to demonstrate the sequence of the plasmid.GDNF protein and its activity were then determined using pcDNA3.1(+)GDNF plasmid transfected eukaryotic cells.Results The results of restriction enzyme and DNA sequencing revealed that GDNF cloning was successful.The recombinant plasmid could express active GDNF protein in the eukaryotic cells.Conclusions Further study on the role of both GDNF and gene therapy is helpful in the treatment Parkinson's disease.