Construction, Expression And Bioactivity Of A Recombinant Fusion Protein Of Consisting Of Liver Cancer Adhension Peptide And GFP

Cancer has become one of the diseases which threaten the health of the human being. With the development of medical technology,tumor targeting is developed.It can kill the tumor cells specially and reduce side effects to the normal cells.Compared with the traditional methods of surgey, radiotherapy and chemotherapy,it has the high selectivity. The key technique in tumor-targeting is to select the vector homing specially to tumor tissues. As a vector , monoclonal antibody has already been used in the research of tumor-targeting therapy.But it also has disadvantages.For example, its high molecular weight,which makes it hard to locate the tumor through vessels and the high pressure of the interstitial solide tumor.The development of technology of phage display peptide provides a very powerful way that can be used to isolate organ and tumor homing peptides.Technology of phage display peptide has many advantages,which makes it a promising method.Our lab has isolated several liver cancer adhension peptides via phage display random peptide library and to verify the efficacy of the peptides in vivo and in vitro.In order to testify the high affinity of the peptides,we used GFP as a living marker to the peptide.The fluorescence of GFP is very stable ,which conquers mch difficulty in using traditional dying.What's more ,its operation is simple to control.We subcloned the gene of the phage peptide A22,A54,A67,B3 and B20 including the gene of domian I into pBV221,which has gfp.Then we constructs the pET-phage LAP-EGFP.The amino acide sequences of A22,A54,A67 have the common motif of PSS or PTT,while the amino acid sequence of B3 has the motif of PS.Futhermore,B20 is negative control.,it hasn't any motif of PSS ,PTT or PS.After partially purified,the fusion proteins were exanmined by cell immunohistochemistry and FCM.We testified the activity of the fusion protein on liver cancer cells.Meanwhile ,we used the normal liver cells as negative control.In all, our study demonstrated that the fusion protein of A22,A54,A67 and B3could bind the liver cancer cells with high affinity.The results also make prepartions for the futher study of targeted therapy.