[Construction of pORF-mIG and activity evaluation of MIG in vitro].
OBJECTIVE
To establish a transformant of monokine induced by interferon-gamma (MIG) with the eukaryotic expression vector for further investigating the efficacy of its use in antitumor gene therapy.


METHODS
The MIG full-length cDNA was amplified from pBLAST-MIG and was cloned into the eukaryote expression vector pORF-mcs, and the resulted recombinant plasmid was named pORF-MIG. The recombinant pORF-MIG was determined with restriction enzyme and sequencing, and then it was transfected into COS-7 cells by Lipfectamin. Expression of the transformant was detected by immunoblot assay, and the bioactivity was assessed by chemotaxis assay.


RESULTS
The restriction analysis and the sequence determination confirmed that the recombinant pORF-MIG contained the MIG full-length cDNA. And the transformants of pORF-MIG expressed the MIG protein which could apparently attract the activated lymphocytes.


CONCLUSIONS
The recombinant pORF-MIG was constructed successfully, and this recombinant eukaryotic expression vector could be used in additional studies on the biological effect of MIG and its use in anti-tumor gene therapy.
Zhang
Ru
R
Laboratory of Biotherapy, Cancer Center, West China Hospital, Sichuan University, Chengdu 610041, China.
Tian
Ling
L
Xiao
Fei
F
Wen
Yan-jun
YJ
li
Jiong
J
Hou
Jian-mei
JM
Zhang
Ling
L
Li
Gang
G
Yao
Bing
B
Chen
Xian-cheng
XC
Mel
Kai
K
D001483:Base Sequence; D054370:Chemokine CXCL9; D019743:Chemokines, CXC / Q000096:biosynthesis* / Q000235:genetics; D005057:Eukaryotic Cells / Q000378:metabolism; D015316:Genetic Therapy; D005822:Genetic Vectors; D006801:Humans; D008969:Molecular Sequence Data; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D014162:Transfection
C080810:CXCL9 protein, human; D054370:Chemokine CXCL9; D019743:Chemokines, CXC; D011994:Recombinant Proteins