Techniques for the single-step amplification of whole genomes have been developed into powerful tools for phylogenetic analyses, epidemiological studies and studies on genome organization. Recently, the bacteriophage phi29 DNA polymerase has been used for the efficient amplification of circular DNA viral genomes without the need of specific primers by a rolling-circle amplification (RCA) mechanism. Various protocols have been applied for detection of novel viruses, for differentiation between circular and linear forms of viral genomes and for generation of infectious genomic clones directly from specimens. Here, we summarize the broad application of the RCA technique to DNA viruses infecting humans, animals and plants.