[Construction of the eukaryote expression vector of human soluble tumor necrosis factor receptor].
OBJECTIVE
Human soluble tumor necrosis factor receptor (sTNFR) can interfere with the biological functions of interleukin-1, which may be appropriate to the treatment of periodontitis. The eukaryote expression vector of the human sTNFR gene must be constructed prior to conducting transgene therapy of periodontitis.


METHODS
Both sTNFR gene and plasmid pcDNA 3.1 (+) DNA were digested with Kpn I and Xho I. After purification, the two fragments were ligated by TakaRa DNA Ligation Kit (Ver 2.0). This recombinant DNA was then transformed into E. Coli Competent Cells JM109 and positive clones were selected on the LB agarose plate containing ampicillin (80 microg/ul).


RESULTS
Six single clones were indentified by double digestion with kpn I and xho I and two fragments with the size of 5.4 kb and 1.0 kb were produced as expected.


CONCLUSIONS
The sTNFR gene was successfully inserted into the eukaryote expression vector plasmid pcDNA 3.1 (+) by recombination technology in vitro.
Xu
Yan
Y
Dept. of Oral Medicine, College of Stomatology, Nanjing Medical University, Nanjing 210029, China.
Zhang
Jin-Cai
JC
Zhang
Yun-Hui
YH
D004926:Escherichia coli; D056890:Eukaryota; D005057:Eukaryotic Cells; D005822:Genetic Vectors; D006801:Humans; D010957:Plasmids; D018124:Receptors, Tumor Necrosis Factor; D047888:Receptors, Tumor Necrosis Factor, Type I; D014409:Tumor Necrosis Factor-alpha
D018124:Receptors, Tumor Necrosis Factor; D047888:Receptors, Tumor Necrosis Factor, Type I; C439524:TNF protein, human; C506235:TNFRSF1A protein, human; D014409:Tumor Necrosis Factor-alpha