A small cryptic plasmid from Ruminobacter amylophilus NIAH-3 possesses functional mobilization properties

The complete nucleotide sequence of a small cryptic plasmid designated pRAO1, from the Gram-negative ruminal bacterium Ruminobacter amylophilus NIAH-3, was determined. The plasmid is a circular DNA molecule, 2140 bp in size, with a GC content of 40%. Computer-assisted analysis identified three open reading frames (ORFs), one of which, ORF3 (347 amino acids), displayed a high degree of amino acid identity with the Mob proteins involved in conjugative mobilization and interplasmid recombination of plasmids from Gram-positive bacteria. We proved the mobilization properties of pRAO1 in the Escherichia coli system using the coresident IncW broad-host-range conjugative plasmid R388. These data demonstrated, for the first time, the mobilization properties of small cryptic plasmids from Gram-negative inhabitants of the rumen.