The role of repetitive DNA sequences in the size variation of Epstein-Barr virus (EBV) nuclear antigens, and the identification of different EBV isolates using RFLP and PCR analysis.

The six Epstein-Barr virus (EBV) nuclear antigen proteins (EBNA-1-6) show characteristic size variations between different virus isolates; this is a feature that has been used to identify the source of virus isolates in epidemiological studies (Ebnotyping). We have now studied the correlation between restriction fragment length polymorphisms (RFLPs) within exons coding for the EBNAs and the molecular masses of the respective proteins. The B95-8 EBV strain was used as the prototype virus. The variation in apparent molecular mass of EBNA-1, -3 and -6 correlated positively with the size of RFLP coding for repeat sequences in these polypeptides. For EBNA-2, no correlation between apparent molecular mass and length of the repetitive sequences was found. The EBNA-4 protein showed virtually no variation in apparent molecular mass and RFLP size across the repeat sequence. Based on the strong correlation between apparent molecular mass and RFLP size for EBNA-6, we developed an EBNA-6 PCR assay that discriminated between different isolates of EBV. This assay offers the advantage of EBV characterization using uncultured material (e.g. throat washings, blood or biopsies), thus avoiding the selection against poorly transforming strains that occurs during establishment of lymphoblastoid cell lines required for Ebnotyping at the protein level.