Chinese Journal of Tissue Engineering Research ›› 2011, Vol. 15 ›› Issue (2): 241-244.doi: 10.3969/j.issn.1673-8225.2011.02.012

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Construction of pcDNA3/BDNF eukaryotic expression vectors

Meng Bu-liang1, Xu Dan2, Liu Jia2, Li Li-yan2, Wang Ting-hua2   

  1. 1Department of Anatomy, 2Institute of Neuroscience, Kunming Medical College, Kunming  650031, Yunnan Province, China
  • Received:2010-07-13 Revised:2010-08-23 Online:2011-01-08 Published:2011-01-08
  • Contact: Wang Ting-hua, Doctor, Professor, Institute of Neuroscience, Kunming Medical College, Kunming 650031, Yunnan Province, China tinghua_neuron@263.net
  • About author:Meng Bu-liang☆, Studying for doctorate, Department of Anatomy, Kunming Medical College, Kunming 650031, Yunnan Province, China mbloso@126.com

Abstract:

BACKGROUND: Brain-derived neurotrophic factor (BDNF) is biomacromolecule, which can not pass through the blood-brain barrier. Now gene therapy is the most promising program to solve route of administration
OBJECTIVE: To construct the eukaryotic expression recombinant plasmid pcDNA3/BDNF.
METHODS: Total RNA was extracted from Sprague Dawley rats using RT-PCR. By gene recombination technique, rat BDNF coding sequence was inserted into eukaryotic expression vector pcDNA3. The fragment of target gene was connected with pcDNA3 vector and transfected into DH5α cells, and recombinant plasmid was verified with restriction enzyme digestion and DNA sequencing.
RESULTS AND CONCLUSION: RT-PCR showed that BDNF product was 749 bp specific segment. By restriction enzyme digestion, the recombinant plasmid consisted to 749 bp and 5 446 bp fragments. The DNA sequence of the 749 bp fragment was identical with rat BDNF cDNA in Gene Bank. The recombinant plasmid pcDNA3/BDNF was constructed successfully.

CLC Number: