Sequence complexity of circular Epstein-Bar virus DNA in transformed cells

Abstract

A simplified procedure, based on several methods previously used to isolate circular DNA molecules from bacteria, was derived for the preparation of covalently closed circular viral DNA molecules from large quantities of lymphocytes transformed by Epstein-Barr virus. The protocol can be applied both to virus nonproducer lines and to lines containing cells activated to virus production. Sufficient amounts o highly purified viral DNA of intracellular origin were obtained from B95-8 and Raji cells to allow direct visual analysis of their sequence complexities after cleavage with EcoRI and separation of fragments by gel electrophoresis. No major differences in complexity were observed between circular DNA and linear virion DNA from B95-8 cells. The fragment patterns observed in this fashion agree well with those detected by conventional blotting and hybridization methods. The procedure can also be used as an analytical method to assay for small amounts of circular Epstein-Barr virus DNA molecules in other transformed cells. In this connection, no circular Epstein-Barr virus DNA was detected in Namalva cells.