The replication pattern of adenovirus DNA in vivo reproduced in vitro

Abstract

Temperature-sensitive mutants of adenovirus type 5 (H5ts125 and H5ts149), which are conditionally inhibited in the initiation of viral DNA synthesis, have been exploited to investigate the possibility of the initiation of replication in a cell-free system. Nuclei were isolated from human KB cells which had been infected with wild-type or mutant adenovirus. More than 90% of the DNA synthesis taking place in such nuclei was virus-specific and the pattern of drug inhibition suggested that the synthesis required DNA polymerase gamma. Nuclei prepared from cells infected with the H5ts125 temperature-sensitive mutant which have been shifted from 33 degrees C to 39.5 degrees C showed a pattern of synthesis in vitro which began at both ends of the viral genome and gradually spread through the rest of the molecule.