IDENTIFICATION AND NUCLEOTIDE-SEQUENCE OF RHIZOBIUM-MELILOTI INSERTION-SEQUENCE ISRM3 - SIMILARITY BETWEEN THE PUTATIVE TRANSPOSASE ENCODED BY ISRM3 AND THOSE ENCODED BY STAPHYLOCOCCUS-AUREUS IS256 AND THIOBACILLUS-FERROOXIDANS IST2

The insertion sequence ISRm3 was discovered simultaneously in different Rhizobium meliloti strains by probing Southern blots of total cellular DNA with P-32-labeled pTA2. This plasmid is indigenous to strain IZ450 and fortuitously contained four copies of ISRm3. By using an internal EcoRI fragment as a specific probe (pRWRm31), homology to ISRm3 was subsequently detected in over 90% of R. meliloti strains tested from different geographical locations around the world. The frequency of stable nonlethal ISRm3 transpositions was estimated to be 4 x 10(-5) per generation per cell in strain SU47 when grown in liquid culture. The entire nucleotide sequence of ISRm3 in R. meliloti 102F70 is 1,298 bp and has 30-bp terminal inverted repeats which are perfectly matched. Analysis of six copies of ISRm3 in two strains showed that a variable number of base pairs (usually eight or nine) were duplicated and formed direct repeats adjacent to the site of insertion. On one DNA strand, ISRm3 contains an open reading frame spanning 93% of its length. Comparison of the putative protein encoded with sequences derived from the EMBL and GenBank databases showed significant similarity between the putative transposases of ISRm3 from R. meliloti, IS256 from Staphylococcus aureus, and IST2 from Thiobacillus ferrooxidans. These insertion sequences appear to be distantly related members of a distinct class.