[Construction of recombinant plasmid human imprinted gene PEG10 and the primary functional identification in transfected cell lines].
OBJECTIVE
To construct a plasmid expressing human imprinted gene PEG10 and to study the effect of overexpression of PEG10 in a stable transfected human normal liver cell line L02 and in non-liver derived cell line 293.


METHODS
Full length cDNA of PEG10 open reading frame 1 was amplified and subcloned into a mammalian expression vector pcDNA3.1hisC. Recombinant plasmid was stably transfected into L02 cells and control cells via Lipofectamine 2000. The expression and the function of PEG10 in L02 cells and control group cells were examined using RT-PCR, Western blot, MTT and TUNEL.


RESULTS
Recombinant plasmid was successfully constructed and confirmed through DNA sequencing and restriction digesting. PEG10 gene accelerated the growth of L02 cells and inhibited their apoptosis but it had no conspicuous effect on the non-liver derived cells.


CONCLUSIONS
The constructed expressing vector pcDNA3.1hisC-PEG10 provides a useful tool for further study on the effects and mechanisms of PEG10. Over-expression of PEG10 may promote L02 cells' proliferation and inhibit their apoptosis, but not in the non-liver derived cell line 293.
Zhang
Qiong
Q
Department of Gastroenterology, Affiliated Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
Xie
Na
N
Wang
Xiao-yan
XY
Chang
Ying
Y
Lin
Yuan-yuan
YY
Lin
Ju-sheng
JS
D051017:Apoptosis Regulatory Proteins; D002460:Cell Line; D004268:DNA-Binding Proteins; D015870:Gene Expression; D005822:Genetic Vectors; D018392:Genomic Imprinting; D006801:Humans; D010957:Plasmids; D011506:Proteins / Q000235:genetics*; D016601:RNA-Binding Proteins; D014162:Transfection
D051017:Apoptosis Regulatory Proteins; D004268:DNA-Binding Proteins; C432965:PEG10 protein, human; D011506:Proteins; D016601:RNA-Binding Proteins