Cloning and sequencing of the genome of spiroplasma virus 4

Spiroplasma virus 4 (Sp V4) has a circular single-stranded DNA. The replicative form (RF) of Sp V4 has been purified from infected cells of Spiroplasma melliferum, strain G1, and cloned in Escherichia coli (HB101) using plasmid pBR328 as the vector. The cloned RF was shown to be infectious by transfection. The Sp V4 RF was randomly subcloned in E. coli (TG1) using the M1 3 mp8 RF as the vector and sequenced by the dideoxy chain termination technique. We found that UGA is probably not a termination codon, but codes for tryptophan. Eight open reading frames, including that for the 65,000-dalton capsid protein, have been detected; they involve all three reading frames.