GENOMIC ANALYSIS OF A TRANSPOSITION-DELETION VARIANT OF ORF VIRUS REVEALS A 3-CENTER-DOT-3KBP REGION OF NONESSENTIAL DNA

Restriction endonuclease analysis of the DNA extracted orf virus strai n NZ2, which had been serially passaged in primary bovine testis cells , revealed a population of variants that had over-grown the wild-type virus. At least three distinct mutant forms were identified in which t he right end of the genome had been duplicated and translocated to the left end, accompanied by deletions of sequences at the left end. Sequ encing of a single variant isolated from the heterogeneous population revealed that recombination had occurred between non-hemologous sequen ces. In this case, 6.6 kb of DNA at the left end of the genome had bee n replaced by 19.3 kb from the right end. The transposition resulted i n the deletion at the left end of 3.3 kb of DNA encoding three genes a nd the terminal sequence of a fourth gene. The three genes completely deleted were a homologue of dUTPase, a gene that encodes a protein con taining ankyrin-like repeats and a homologue of the 5K gene of the vac cinia virus WR strain. Experimental inoculation of sheep showed that t he genes are also non-essential in vivo, but that the size of the lesi on was reduced, compared with that induced by the wild-type, and resol ved more rapidly.