Cloning of chicken anemia virus vp3 gene and apoptosis inductive effect of vp3 gene in vitro.
Using PCR technique, the vp3 gene of chicken anemia virus (CAV) was cloned into the eukaryotic expression vector pcDNA3 to construct a recombinant pcDNA-vp3. Restriction enzyme digestion and sequencing analysis revealed that CAV vp3 gene was correctly inserted into the blank vector pcDNA3. After LipofectAMINE-mediated transfection in vitro with pcDNA-vp3 and pcDNA3 respectively, the total mRNA was extracted from liver carcinoma cell lines HepG2 and diploid cell line L-02, and RT-PCR was performed afterward. The results of RT-PCR suggested that vp3 gene was expressed in these two cell lines. At the same time, using in situ apoptotic detection assay, TUNEL kits, the apoptotic cells were found in pcDNA-vp3 transfected HepG2, but not in mock transfected cell lines. VP3 could induce cell death by apoptosis in cancer cell lines, but not in diploid cell lines. All the results indicated that CAV vp3 gene, a potential therapeutic agents, has the potential of being used for cancer treatment.
Sun
Jun
J
Department of Biochemistry and Molecular Biology, School of Basic Medical Sciences, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030.
Wang
Yuzhe
Y
Zong
Yiqiang
Y
Qu
Shen
S
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D036022:Capsid Proteins / Q000235:genetics*; D006528:Carcinoma, Hepatocellular / Q000473:pathology; D045744:Cell Line, Tumor; D017727:Chicken anemia virus; D002645:Chickens; D003001:Cloning, Molecular; D005057:Eukaryotic Cells; D005822:Genetic Vectors; D006801:Humans; D008055:Lipids; D008113:Liver Neoplasms / Q000473:pathology*; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000494:pharmacology; D017422:Sequence Analysis, DNA; D014162:Transfection; D014764:Viral Proteins / Q000235:genetics
D036022:Capsid Proteins; D008055:Lipids; C086724:Lipofectamine; D011994:Recombinant Proteins; C091717:VP3 protein, Chicken anemia virus; D014764:Viral Proteins