INVESTIGATION OF THE TRANSFECTION CAPABILITY OF CLONED TANDEMLY-REPEATED CHICKEN ANEMIA VIRUS-DNA FRAGMENTS

Chicken anaemia virus (CAV) is an icosahedral virus, 25 nm in diameter , which, on the basis of its circular single-stranded DNA genome, has recently been classified in the family, Circoviridae. We have investig ated whether infectious, monomeric CAV DNA from recombinant plasmids c ontaining tandemly-repeated CAV replicative form (RF) DNAs, following transfection, was generated by homologous recombination or a replicati onal release mechanism involving rolling circle replication (RCR) of D NA. Experiments designed to locate the virus strand origin of RCR and/ or sites of recombination were performed by sequence analyses of hybri d viruses generated after transfection with cloned tandemly-repeated R Fs specified by the sequence-distinct Cux-1 and 26P4 isolates. Positiv e transfection results obtained from 2 recombinant plasmid constructs were shown to have resulted from homologous recombination occurring at different sites within the RF sequence. Three of 5 hybrid viruses ana lysed were ''circularised'' within the same 105 bp sequence, that cont ains four 19 bp repeats and with which promoter/enhancer activity has been associated. This region may represent a novel origin or recombina tion hot-spot within the CAV genome. A distinctive cruciform-loop stru cture within the non-coding region was shown to contain an S1 nuclease -sensitive site, detected in CAV RF and in recombinant plasmids contai ning RF inserts.