While transposon mutagenesis has been successfully used for @entity9463 spp. to disrupt and determine non-essential genes, previous attempts with @entity12726 spp. have been unsuccessful. Using a @entity968 -transformation enhancing protocol, we were able to transform three separate serovars of @entity24757 with a Tn4001-based mini-transposon plasmid containing a @entity801 resistance selection marker. Despite the large degree of homology between @entity24757 and @entity12726 , all attempts to transform the latter in parallel failed, with the exception of a single clinical @entity12726 isolate. PCR probing and sequencing were used to confirm transposon insertion into the bacterial genome and identify disrupted genes. @entity4383 serovar 3 consistently resulted in transfer only of sequence between the mini-transposon inverted repeats, but some strains showed additional sequence transfer. Transposon insertion occurred randomly in the genome resulting in unique disruption of genes UU047, UU390, UU440, UU450, UU520, UU526, UU582 for single clones from a panel of screened clones. An intergenic insertion between genes UU187 and UU188 was also characterised. Two phenotypic alterations were observed in the mutated strains: Disruption of a DEAD-box RNA helicase (UU582) altered growth kinetics, while the @entity12726 lost resistance to serum attack coincident with disruption of gene UUR10_137 and loss of expression of a 41kDa protein. Transposon mutagenesis was used successfully to insert single copies of a mini-transposon into the genome and disrupt genes leading to phenotypic changes in @entity24757 strains. This method can now be used to deliver exogenous genes for expression and determine essential genes for @entity24757 replication in culture and experimental models.

Random insertion and gene disruption via transposon mutagenesis of XXXX using a mini-transposon plasmid.

@entity9463 :: ('MESH:D009175', 'Disease') :: ['Mycoplasma'] @entity801 :: ('MESH:D005839', 'Chemical') :: ['gentamicin'] @entity24757 :: ('134821', 'Species') :: ['Ureaplasma parvum'] @entity4383 :: ('MESH:D020518', 'Disease') :: ['Transformation of prototype'] @entity12726 :: ('2130', 'Species') :: ['U. urealyticum strain', 'Ureaplasma urealyticum', 'Ureaplasma', 'U. urealyticum'] @entity968 :: ('MESH:D011092', 'Chemical') :: ['polyethylene glycol']
@entity24757 :: (134821,Species) :: ['Ureaplasma parvum']
