食管癌组织中HPV16型L2线性抗原基因的克隆及原核表达

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Cloning and expression of HPV16 L2 DNA from esophageal carcinoma in E. coli

AIM To acquire the HPV16 L_2 conservative antigen DNA from tissues of esophageal carcinoma infected with human papillomavirus by PCR using special primers, and to sequence the subjective DNA by cloning into vector of pGEM-3zf(-) and express L_2 conservative antigen DNA in E. coli, and assay its immunoreaction with polyclone antibody or patients sera. METHODS The conservative antigen DNA segment was ascertained by comparing the L_2 sequence of human papillomavirus 16 through BLAST 2.0 on NCBI, then using specific primers, the subjective DNA segment was acquired by PCR from tissues of esophageal carcinoma infected with human papillomavirus. After that it was cloned into vector of pGEM-3zf(-) and sequenced, and inserted into an expression vector and expressed in E. coli. The antigen activity of expressed product was detected by Western-blot. RESULTS The desired DNA of HPV16 L_2 conservative antigen can be acquired by PCR using special primer from the tissues of esophageal carcinoma infected with human papillomaviruso the acquired DNA fragment can be cloned into vector of pGEM-3zf (-) and sequenced. The DNA sequence from two cases was highly conserved. It can also be cloned into an expression vector and expressed actively in E. coli. CONCLUSION The HPV16 L_2 conservative antigen DNA can be acquired from tissue of esophageal carcinoma Infected with human papillomavirus by PCR, and there is high conservation in the tissue of esophageal carcinoma infected with human papillomavirus. The product expressed in E. coil was active, which may provide a cheap and convenient method for epidemiology and presentiment of the occurence of cancer related with human papillomavirus.