Establishment and Application of a Method for High-Risk Human Papillomavirus Genotyping in Cervical Cancer Tissue.

Abstract：

BACKGROUND:Persistent high-risk HPV infection is a major cause of cervical cancer and E6/E7 genes and the Li gene in the HPV genome are key targets to detect high-risk HPV. This study aims to explore the relationship between cervical lesions and E6/7 by establishing a polymerase chain reaction (PCR) to detect multiplex genes based on HPV EE7 genes. It is hoped that such methods will provide a more reliable method for clinical screening and the prevention of cervical cancer. METHODS:Based on alignment, specific primers were designed for HPV E6/E7 genes, the sequences of which came from five5 high-risk papillomaviruses that are common in China. This enabled an E6/E7 gene detection method based on multiplex PCR to be established. E6/E7 and Li gene testing were then performed on 65 cervical cancer tissue samples. The gene copy number of HPV E6/E7 genes and the Li gene were detected from different classifications by real-time fluorescence quantitative PCR. RESULTS:Out of the 65 cervical cancer tissue samples, 47 (72.31%) showed positive results in E6/E7 multiplex PCR, 21 (32.31%) showed positive results in the Ll gene PCR, and out of the 219 cervical exfoliate cell samples, 56 (25.57%) showed positive results in E6/E7 multiplex PCR, 21 (13.24%) showed positive results in the L1 gene PCR. There were significant differences (p < 0.05) between these two test results. Fluorescent quantitative PCR showed that the ratio of gene copy number of L1 genes and E6/E7 genes was below 1 (p < 0.05) in cervical cancer tissue, in which both the Li and E6/E7 genes coexist. CONCLUSIONS:The established HPV multiplex PCR assay based on the design of E6/E7 gene is a specific and sensitive method for the detection and genotype of five high-risk HPVs.