Analysis of the DNA produced during infection of monkey kidney cells by adenosatellite virus type 4 and adenovirus SV15.

Abstract

SUMMARY Radioisotope labelling, autoradiography, electron microscopy and DNA-DNA hybridization have been used to analyse the DNA species produced in a stable line of monkey kidney cells after infection with defective adeno-associated satellite virus (ASV), in presence of and without simian helper adenovirus (SV 15). Extraction of DNA by the Hirt (1967) procedure and CsCl equilibrium density gradient sedimentation indicated that a newly synthesized low mol. wt. DNA was readily isolated from cells infected with the defective satellite virus alone, within 6 h after inoculation. Sedimentation analyses of this DNA through neutral sucrose gradients showed that it was heterogeneous, with a sedimentation coefficient in the range 11 to 23 S. The DNA was non-infectious when titrated with helper adenovirus. Sedimentation analyses of DNA extracted from cultures infected with satellite virus and helper adenovirus showed linear 32 S adenovirus DNA and a 26 S linear DNA which may represent a replicative intermediate of satellite virus. Electron microscopy showed that the latter DNA was linear with marked propensity for clumping and concatenation. Molecules with side chains were seen. Autoradiography showed that newly synthesized DNA isolated from cells infected with satellite alone was nuclear in origin and DNA-DNA hybridization indicated that the synthesized DNA not extracted by the Hirt procedure was cellular rather than virus DNA.