Cloning of human papilloma virus genomic DNAs and analysis of homologous polynucleotide sequences

Abstract

The complete DNA genomes of 4 distinct human papilloma viruses (human papilloma virus subtype 1a [HPV-1a], HPV-1b, HPV-2a and HPV-4) were molecularly cloned in Escherichia coli, using the certified plasmid vector pBR322. The restriction endonuclease patterns of the cloned HPV-1a and HPV-1b DNA were similar to those already published for uncloned DNA. Physical maps were constructed for HPV-2A DNA and HPV-4 DNA, since these viral DNA had not been previously mapped. By using the cloned DNA, the genomes of HPV-1a, HPV-2 and HPV-4 were analyzed for nucleotide sequence homology. Under standard hybridization conditions (Tm [melting temperature] = -28.degree. C), no homology was detectable among the genomes of these papilloma viruses, in agreement with previous reports. Under less stringent conditions (i.e., Tm = -50.degree. C), stable DNA hybrids could be detected between these viral DNA, indicating homologous segments in the genomes with approximately 30% base mismatch. By using specific DNA fragments immobilized on nitrocellulose filters, these regions of homology were mapped. Hybridization experiments between radiolabeled bovine papilloma virus type 1 (BPV-1) DNA and the unlabeled HPV-1a, HPV-2a or HPV-4 DNA restriction fragments under low-stringency conditions indicated that the regions of homology among the HPV DNA are also conserved in the BPV-1 genome with approximately the same degree of base mismatch.