Application of TDI-FP for Analysis of A647G Variation in HPV 16 E7 Gene From Cervical Cancer Patients / 生物化学与生物物理进展

ABSTRACT

Template direct dye-terminator incorporation with fluorescence-polarization (TDI-FP assay) is a technology for genotyping single nucleotide polymorphisms (SNPs). To apply this method in analyses of A647G variation in human papillomavirus (HPV) 16 E7 gene from HPV 16-positive cervical tissues, a total of 91 and 49 HPV 16-positive DNA samples obtained from women with cervical cancer and normal/inflamed cervices living in Shaanxi in northwest China were subjected to the partial E7 gene PCR with nucleotide (nt) 647 in the products. Then, the oligonucleotide probe designed to anneal immediately to nt 647 was hybridized to the template within the PCR amplicons, and extended specifically by TAMRA-ddTTP or R110-ddCTP directed by the base at nt 647. The increasing FP values were read and the base at nt 647 was identified. The prevalence of nt 647 A→G was 35.71% (50/140). The variation 647G detected in 42.86% (39/91) of women with cervical cancer was significantly higher than 22.45% (11/49) detected in those with normal/inflamed cervices (x2 = 5.778, P = 0.016). The odds ratio (OR) between these two groups was 2.59 (95% confidence interval=l.17～5.71). The results demonstrate that TDI-FP method can be potentially applied in analysis of interest point mutations in HPVs. The incidence and risk implication of HPV 16 A647G variant infection in Shaanxi, China, displays significant geographic difference from other areas. The HPV 16 with E7 gene A647G point mutation appears to have a higher risk for invasive cervical cancer in women living in Shaanxi.