Identification and characterization of IS1302, a novel insertion element from Wolinella succinogenes belonging to the IS3 family

Abstract

A new insertion sequence (IS) designated IS1302 was identified in Wolinella succinogenes. IS1302 is 1,306 bp in size with 36-bp imperfect terminal inverted repeats. It contains only one open reading frame (tnpA), which encodes a putative transposase whose sequence is similar to that of transposases of various IS elements of the IS3 family. IS1302 was identified in the genome of a W. succinogenes fumarate reductase deletion mutant in which the frd operon had been replaced by the kan gene. The insertion of IS1302 occurred when the mutant was propagated in the presence of a high concentration of kanamycin. Two different target sites of IS1302 were found immediately upstream of the kan gene, where the insertion of IS1302 resulted in a duplication of 3 bp of the target DNA. Upon insertion of IS1302, new possible promoter structures of the kan gene were created, which might lead to a stimulated transcription of the kan gene and result in a selective advantage of cells containing IS1302 at one of the two target sites. Southern blot analysis suggested the presence of at least 13 copies of IS1302 in the genome of W. succinogenes. This is the first IS element discovered in W. succinogenes.