Transposition of Tn 916 in the four replicons of the Butyrivibrio proteoclasticus B316.

The rumen bacterium Butyrivibrio proteoclasticus B316 has a 4.4-Mb genome composed of four replicons (approximately 3.55 Mb, 361, 302 and 186 kb). Mutagenesis of B316 was performed with the broad host-range conjugative transposon Tn 916 to screen for functionally important characteristics. The insertion sites of 123 mutants containing a single copy of Tn 916 were identified and corresponded to 53 different insertion points, of which 18 (34.0%), representing 39 mutants (31.7%), were in ORFs and 12 were where transposition occurred in both directions (top and bottom DNA strand). Up to eight mutants from several independent conjugation experiments were found to have the same integration site. Although transposition occurred in all four replicons, the number of specific insertion sites, transposition frequency and the average intertransposon distance between insertions varied between the four replicons. In silico analysis of the 53 insertion sites was used to model a target consensus sequence for Tn 916 integration into B316. A search of the B316 genome using the modelled target consensus sequence (up to two mismatches) identified 39 theoretical Tn 916 insertion sites (19 coding, 20 noncoding), of which nine corresponded to Tn 916 insertions identified in B316 mutants during our conjugation experiments. [ABSTRACT FROM AUTHOR]