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Expression, Isolation, and Purification of Soluble and Insoluble Biotinylated Proteins for Nerve Tissue Regeneration
Published on: January 22, 2014
[Prokaryotic expression and activity identification of gene recombinant protein of brain-derivedneurotrophic factor
Jia Chen1, Xiaomin Liang1, Zhiqiang Xu1
1Department of Neurology, Daping Hospital-Field Surgery Institute, Third Military Medical University, Chongqing 400042, China.
Objective:
To generate the gene recombinant protein of brain-derived neurotrophic factor precursor (proBDNF) in prokaryotic cells and investigate its biological activity.
Methods:
Rat-derived cDNA of proBDNF with point mutation was amplified by polymerase chain reaction (PCR) and cloned into plasmid pET-28a for expression in E. coli BL21. Western blot was used to identify the product and DAPI performed to test its effect on apoptosis of PC12 cells.
Results:
PCR product of recombinant gene was successfully expressed in E. coli. And the product agreed with the target protein in molecular weight and showed reactivity with its specific antibody. Apoptosis of PC12 cells was induced by a certain concentration of recombinant proBDNF.
Conclusion:
The prokaryotic expression vector has been successfully constructed for recombinant gene of proBDNF. And the product has biological toxicity and it may induce the apoptosis of PC12 cells.
