[Expression of HCV NS5A gene in HepG2 cell and the effect of the NS5A expression on PI3K in vitro]

To reveal the influence of HCV-NS5A on PI3K, we probe into the relationship between NS5A and PI3K in vitro.

Full length NS5A gene of HCV was amplified by PCR, using the plasmid containing HCV full-length open reading frame (ORF) as template, and cloned into the eukaryotic expressing plasmid pcDNA3.0(-) by DNA recombination technique. The recombinant vector was identified by digestion with restriction enzymes and polymerase chain reaction and by directly sequencing. Then both the recombinant vector pcDNA3.0(-)-NS5A and the control vector pcDNA3.0(-) were transfected HepG2 cell using Lipofectamin2000.

Expressing NS5A was proven by RT-PCR and Western blot analysis in transfected HepG2 cell. Further more we examined the PI3K protein expressed in the HepG2 cell expressing recombinant NS5A.

NS5A can activate PI3K in vitro and its signal cascade pathway.