Cloning, expression and biological activity analysis of rat GFRα1 gene

To obtain recombinant glial cell line-derived neurotrophic factor receptor alphal (GFRalpha1) and study its biological activity, the cDNA encoding the mature rat GFRalpha1 was isolated using RT-PCR with total RNA extracted from newborn SD-rat hippocampus tissue. The expression plasmid pET-GFRalpha1 was constructed by inserting GFRalpha1 cDNA into plasmid pET-28a ( +) containing T7 promoter and transformed into E. colt BL21 (DE3). An expression strain BLGFRalpha1 was selected. SDS-PAGE analysis revealed that the rat GFRalpha1 protein was highly expressed and accumulated up to 21.5% of the total bacterial proteins in the form of inclusion body after the induction. By Ni2+ chelation affinity chromatography, up to 90% GFRalpha1 protein was purified. Purified and refolded GFRalpha1 protein could significantly mediate the ability of GDNF to promote the survival and induce the differentiation of PC12 cells.