Rescue of defective SV40 from a transformed mouse 3T3 cell line: Selection of a specific defective

Small-plaque type SV40 yields various defectives containing deletions, insertions, and substitutions. To select a specific defective lacking a part of the late genes, we attempted to rescue a defective mutant from a transformed mouse cell line. A triple heterokaryon culture was prepared which consisted of monkey cells, 3T3 cells transformed by defective SV40, and 3T3 cells transformed by large-plaque type SV40 that yields no defective virus having the capacity to direct T antigen synthesis. Virus recovered from the heterokaryon culture was propagated in monkey cells, and defective light virions were concentrated by CsCl equilibrium centrifugation. The virions and their DNA thus obtained were found to direct T antigen synthesis but not synthesis of virion antigen. Heteroduplex DNA molecules presumably consisting of a complete and a defective strand had mostly two single-stranded loops. The rescued defective SV40 genome appeared to have a deletion and an insertion at two separate and fixed sites.