Construction of anEnterococcus faecalisTn917-Mediated-Gene-Disruption Library Offers Insight into Tn917Insertion Patterns

Abstract

Sequencing the insertion sites of 8,865 Tn917insertions inEnterococcus faecalisstrain OG1RF identified a hot spot in the replication terminus region corresponding to 6% of the genome where 65% of the transposons had inserted. InE. faecalis, Tn917preferentially inserted at a 29-bp consensus sequence centered on TATAA, a 5-bp sequence that is duplicated during insertion. The regional insertion site preference at the chromosome terminus was not observed in another low-G+C gram-positive bacterium,Listeria monocytogenes, although the consensus insertion sequence was the same. The 8,865 Tn917insertion sites sequenced inE. faecaliscorresponded to only ∼610 different open reading frames, far fewer than the predicted number of 2,400, assuming random insertion. There was no significant preference in orientation of the Tn917insertions with either transcription or replication. Even though OG1RF has a smaller genome than strain V583 (2.8 Mb versus 3.2 Mb), the onlyE. faecalisstrain whose sequence is in the public domain, over 10% of the Tn917insertions appear to be in a OG1RF-specific sequence, suggesting that there are significant genomic differences amongE. faecalisstrains.