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Published on: January 19, 2011
[Construction of the recombinant plasmid of pcDNA3.0-RKIP and expression in PC12 cells]
Tao Lin1, Hong-yan Zuo, De-wen Wang
1Institute of Radiation Medicine, Academy of Military Medical Sciences, Academy of Military Medical Sciences, Beijing 100850, China. shanialin@126.com
Aim:
To construct the eukaryotic expression vectors of RKIP plasmid and detect its expression in PC12 cells.
Methods:
The coding sequence of RKIP was generated by nested-PCR using total RNA extracted from the root ganglion neurons of rats. RKIP gene was cloned into the eukaryotic expression vector pcDNA3.0. After restriction enzyme analysis and sequence identification, the recombinant plasmid was transfected into PC12 cells with non-liposome mediated method by Vigofect. The expression of RKIP was detected by Western blot.
Results:
The results of enzyme analysis and sequencing both identified DNA sequence of recombinant plasmid pcDNA3.0-RKIP correctly. The expression of RKIP increased obviously after transfection into PC12 cells.
Conclusion:
The eukaryotic expression plasmid of pcDNA3.0-RKIP was constructed successfully and it can be sustainly expressed in PC12 cells. This provides experimental basis for further study on the neurological function of RKIP.
