Objective To study the relationship of HPV pseudo-neutralizing titers detected by two different reporter genes: Zoanthus sp. green fluorescent protein (ZsGreen) and secreted alkaline phosphatase (SEAP) , and the relationship between HPV the pseudovirus-neutralizing antibody titer and the antibody titer determined by ELISA method. Methods The plasmids with expression cassettes of the HPV capsid protein L1 and L2 genes after codon optimization and the plasmid with reporter gene (ZsGreen or SEAP) were co-transfected into 293FT cells. The cell lysate supernatants were collected after 48 h culture, then the pseudovirus was purified through POROS column chromatography from the supernatants. After the titer of pseudovirus bulk were measured, HPV-16 and HPV-18 pseudovirus-neutralization assays were carried out for determining the titer of sera collected from immunized mice with HPV candidate vaccine and Gardasil HPV vaccine. Results In statistical analysis, the two reporter gene systems for the detection of the pseudovirus neutralizing antibody titer are highly relevant to each other (Spearman coefficient; r = 0. 760). And their neutralizing antibody titers bear a high degree of correlation with the antibody titer (Spearman coefficient: r= 0.577 and r =0. 741). Conclusion ZsGreen and SEAP pseudovirus neutralizing antibody titers are highly relevant to each other. The neutralizing antibody and the antibody titer are also relevant. These results reveal some mechanism of HPV vaccines to prevent the virus from invading the host cells, and are absolutely useful in the protection efficiency evaluation of the HPV-16 and HPV-18 candidate vaccines.

Objective To construct the recombinant X-linked inhibitor of apoptosis protein(XIAP) gene 3′untranslational region (3′UTR)-luciferase reporter vector ,and analyze the microRNA(miRNA) which possibly regulate the expression of XIAP gene . Methods Polymerase chain reaction (PCR) was employed to amplify X IA P-3′UTR sequences from human cDNA ,in which luciferase reporter vector pGL3-Ctrl was inserted ,and the recombinant vector pGL3-Ctrl/XIAP was gained .Target Scan 6 .2 soft-ware was adopted to predict the miRNA which possibly combined with the X IA P-3′UTR .pGL3-Ctrl/XIAP recombinant plasmids and the miRNA were co-transfected into A549 cells ,and the X IA P-3′UTR-luciferase activity was measured .Results Confirmed by digestion and DNA sequencing ,the X IA P-3′UTR-luciferase reporter recombinant was successfully constructed .Prediction of miRNA target sites indicated that X IA P gene may be the target of miR-200b ,miR-200c and miR-429 .Compared with miRNA mim-ic ctrl group ,miR-200b ,miR-200c and miR-429 significantly reduced the luciferase activity of pGL 3-Ctrl/XIAP with statistically significant difference(P<0 .05) .Conclusion X IA P-3′UTR-luciferase reporter vector is successfully constructed .miR-200b ,miR-200c and miR-429 can obviously decrease the luciferase activity .