ANALYSIS OF DELETION OF THE INTEGRATED HUMAN PAPILLOMAVIRUS-16 SEQUENCE IN CERVICAL-CANCER - A RAPID MULTIPLEX POLYMERASE CHAIN-REACTION APPROACH

A protocol for a rapid physical mapping of the integrated type 16 human papillomavirus (HPV16) sequences in biospied and paraffin-embedded archival cervical cancer samples is described. The procedure involves the use of an anchor primer and a mixture of indicator primers in a multiplex polymerase chain reaction (PCR). A minimal conserved region of viral integration of 2,745 bp in length has been mapped between nucleotide (nt) 6102-941, containing the entire regulatory region and the E6 and E7 open reading frames (ORFs). A general deletion domain of 1,465 bp in the integrated viral genome has been defined between nt 1417-2881,covering most of the El ORF at the 3'-half and 60 bp at the 5' terminus of the E2 ORF. This common deleted sequence contains an ATPase active domain speculated to be associated with a DNA helicase function essential for the viral replication, and it also falls within the actively spliced E1-E2 segment of the primary RNA transcripts. Detection of the loss of the 3'-half of the El ORF would be an ideal marker for PCR-based rapid determination of HPV integration in cervical cancer cells. (C) 1994 Wiley-Liss, Inc.