The Experimental Study On Recombinant Expression And Anti-tumor Effect Of ING4

Objective:To construct the recombined adenovines vector of mING4 and explore the effect of ING4 on mSMMC7721 cells.Methods: The pAdTrack-CMV- mING4 was constructed with pcDAN3.0- mING4 recombined plasmid by PCR as template,enzyme digestion and ligation.The pAdTrack-CMV- mING4 lineared by PmeI was co-transducted into BJ5183 with pAdeasy-I.Analysis of PCR and DNA sequencing was carried out to demonstrate the sequence of the plasmid.The pAdeasy-1-pAdTrack-CMV- mING4 recombined adenovirus vector was leared with PacI and then transfected into QBI-293A cells.The mING4 recombined adenovirus was obtained and was used to infect SMMC7721 cells.Identification of mING4 mRNA by RT-PCR was carried out to demonstrate the expression of mING4 in SMMC7721cells.The effect of mING4 on SMMC7721 was studied by MTT,flow cytometry and western blot.Results: Analysis by restricting enzyme digestion and PCR of pAdTrack-CMV- mING4 recombiant plasmid showed results were about 750bp,DNA sequencing revealed that mING4 cloning were successful.Cell cycle and apoptosis were examined by flow cytometry after infection of SMMC7721 cells with pAdeasy-1-pAdTrack-CMV-mING4(Ad- mING4). The inhibition of mING4 on growth of SMMC7721 cells was tested by MTT.We found that mING4 could inhibit the growth and enhance apoptosis in SMMC7721cells.