Mycoplasma virus L2 is a quasi-spherical enveloped virion containing circular double-stranded DNA. L2 infection of Acholeplasma laidlawii host cells leads to a noncytocidal productive infection cycle followed by establishment of lysogeny in all (or most) infected cells, with viral DNA integrated into the host cell genome. The L2 genome has been sequenced and analyzed. L2 DNA is 11,965-bp long and contains 15 open reading frames (ORFs). One of these, ORF13*, has its start codon within and in the same reading frame as ORF13. The ORFs are clustered in four groups separated by noncoding intergenic regions, suggesting that gene expression involves transcription of genes in a cluster into polycistronic mRNA and translation of these genes via translational coupling or reinitiation. Fifteen L2 start codon sites have been defined and resemble those of eubacteria. The N-terminal sequences of two ORFs appear to be signal peptides, and the gene product of one of these may be an L2 virion integral membrane protein. The ORF 5 product has been tentatively identified as an integrase, based on its sequence similarity to site-specific recombinases. The putative attP integration site has been mapped to an intergenic region, 280-bp downstream from ORF 5. Two putative DNA replication ori sites have been mapped. Each is in an intergenic region and contains a DnaA-box bounded by A + T-rich 6-mer repeats.