Replacing CTL epitope of recombinant HBsAg by SOEing PCR
- VernacularTitle:利用重叠PCR实现乙肝表面抗原CTL表位的替换
- Author:
Wenjing WANG
;
Chengyao LI
;
Mingsong LI
- Publication Type:Journal Article
- Keywords:
epitope shift;
glypican-3;
CTL epitope;
SOEing PCR
- From:Journal of Third Military Medical University
2003;0(07):-
- CountryChina
- Language:Chinese
-
Abstract:
Objective To replace the endogenous CTL epitope LLD in HBsAg with EYILSLEEL of glypican(GPC3)in order to prepare a GPC3-HBsAg multiple peptides vaccine for HBV infection.Methods The HBsAg/GPC3 DNA was amplified by gene splicing by overlap extension(SOEing)PCR from pcHBsAg plasmid and then inserted into pBSSK+ vector to construct a pBSSK/GPC3 vector.The vector was then identified by PCR,double digesting and sequencing.The fragment encoding GPC3 CTL epitope EYILSLEEL was obtained by double digesting the plasmid pBSSK/GPC3 and then inserted into pcDNA3.1+ vector.Results Sequencing and restrict endonulease digestion indicated that eukaryotic expression vector pcDNA-HBsAg/GPC3 was constructed successfully.Conclusion The endogenous CTL epitope LLD of HBsAg is replaced by the EYILSLEEL,and an eukaryotic expression vector pcDNA-HBsAg/GPC3 is constructed.
