[Construction of recombinant adenovirus vector carrying human TIMP-1 cDNA and its expression in vitro].
The full-length cDNA of hTIMP-1 was obtained from a surgical patient with HCC by the method of RT-PCR. Then it was cloned into the adenoviral shuttle plasmid pAdTrack-CMV, and subsequently cotransformed into competent BJ5183 cells with the adenoviral backbone plasmid pAdEasy-1. Thereupon, a recombinant adenoviral plasmid containing full-length cDNA of hTIMP-1 was generated by homologous recombination in E. coli. The adenoviruses (AdhTIMP-1) were packaged and amplified in adenoviral packaging cells HEK 293. Then the viral titer was checked by green fluorescent protein (GFP), and the expression of hTIMP-1 was detected by the techniques of Western blot and RT-PCR. The recombinant adenovirus vector carrying human TIMP-1 was successfully constructed and expressed in vitro and may pave the way for further application in liver gene therapy.
Xia
Dong
D
Department of General Surgery, West China Hospital of Sichuan University, Chengdu 610041, China.
Yan
Lünan
L
Xu
Liang
L
Tong
Yu
Y
Zuo
Huaiquan
H
Zhao
Lanying
L
D000256:Adenoviridae / Q000235:genetics / Q000378:metabolism*; D006528:Carcinoma, Hepatocellular / Q000378:metabolism; D002460:Cell Line; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000235:genetics; D005109:Extracellular Matrix / Q000378:metabolism*; D005822:Genetic Vectors; D006801:Humans; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis* / Q000235:genetics; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000096:biosynthesis* / Q000235:genetics
D018076:DNA, Complementary; D011993:Recombinant Fusion Proteins; D019715:Tissue Inhibitor of Metalloproteinase-1