The Double Polymerase Chain Reaction with Consensus Primers Permits Rapid and Sensitive Detection of Genital Human Papillomavirus Oncogenes

概要

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We have developed a sensitive procedure for the detection of relatively low copy numbers of multiple genital human papillomaviruses (HPVs) using the polymerase chain reaction (PCR). HPV DNAs were detected by agarose gel electrophoresis and ethidium bromide staining after 2 rounds of PCR amplification (double PCR) with outer and inner consensus primer pairs for HPV-6, 11, 16, 18, 31, 33, 52, and 58. The detection limit of this method (i. e., 10?? copy of HPV DNA per cell in 1 μg cell DNA) was sufficient for analysis of cervical intraepithelial neoplasia (CIN) specimens. Overall prevalence rate of HPV was 100% in 20 cases of CIN specimens. HPV typing by restriction enzyme analysis revealed that HPV-16 sequence was present in 11 cases, HPV-18 in 1 case, HPV-31 in 4 cases, HPV-33 in 1 case, HPV-52 in 2 cases, HPV-58 in 3 cases, and an unidentified type(s) in 3 cases. There were 4 cases of mixed infections. This procedure obviates the use of hybridization- based for-mat for identification of at least 8 types of HPV sequences present in a small fraction of cells within a heterogeneous population.