Progress towards an inducible, replication-proficient transposon delivery vector for Chlamydia trachomatis

BackgroundGenetic systems have been developed forChlamydiabut the extremely low transformation frequency remains a significant bottleneck. Our goal is to develop self-replicating transposon delivery vector forC. trachomatiswhich can be expanded prior transposase induction.MethodsWe madeE. coli/C. trachomatisshuttle vectors bearing theHimar1C9 under control of thetetpromoter and novel rearrangement theHimar1transposon with ?-lactamase gene. Activity was monitored by immunoblot DNA sequencing.ResultsWe constructed pSW2-mCh-C9, aC. trachomatisplasmid designed act as carrying both undertetpromoter its transposon. However, we were unable recover this plasmid inC. trachomatisfollowing multiple attempts at transformation.Therefore, assembled two new deletion plasmids pSW2-mCh-C9-?Tpon only (undertetpromoter control) sister (same sequence backbone) pSW2-mCh-C9-?Tpase cognate We demonstrated that biological components make up are active inE. coli. Both these could independently recovered trachomatis.We attempted perform lateral gene transfer mixed infection withC. trachomatisstrains bearingpSW2-mCh-C9-?Tpon pSW2-RSGFP-Tpon(a green fluorescent version ofpSW2-mCh-C9-?Tpase). Despite success in achieving infections, it not possible progeny versions plasmids.ConclusionsWe pSW2-mCh-C9 trachomatiscarrying control. Whilst transformed intoE. coliit cannot trachomatis. Based on selected deletions phenotypic analyses conclude level expression from thetetinducible promoter responsible for premature transposition hence loss early process.