title: Dendrimer-enabled DNA delivery and transformation of Chlamydia pneumoniae.
abstract: The chlamydiae are important human pathogens. Lack of a genetic manipulation system has impeded understanding of the molecular bases of virulence for these bacteria. We developed a dendrimer-enabled system for transformation of chlamydiae and used it to characterize the effects of inserting the C. trachomatis plasmid into C. pneumoniae, which lacks any plasmids. The plasmid was cloned into modified yeast vector pEG(KG) and the clone complexed to polyamidoamine dendrimers, producing 50-100 nm spherical particles. HEp-2 cell cultures were infected with C. pneumoniae strain AR-39. Twenty-four hours later, medium was replaced for 3 hours with dendrimer-plasmid complexes, then removed and the medium replaced. Cultures were harvested at various times post-transformation. Real-time PCR and RT-PCR of nucleic acids from transformed cultures demonstrated plasmid replication and gene expression. The cloned plasmid was replicated and expressed in transformants over 5 passages. This system will allow study of chlamydial gene function, allowing development of novel dendrimer-based therapies.
  
  
  BACKGROUND
  This team of investigators developed a dendrimer-enabled system for transformation of chlamydiae and successfully utilized it to characterize the effects of inserting the C. trachomatis plasmid into C. pneumonia. This system will allow study of chlamydial gene function, allowing development of novel dendrimer-based therapies.
authors: Department of Immunology and Microbiology, Wayne State University School of Medicine, Detroit, MI, USA.
authors: Hervé C
authors: HC
authors: Gérard
authors: Manoj K
authors: MK
authors: Mishra
authors: Guangzhao
authors: G
authors: Mao
authors: Sunxi
authors: S
authors: Wang
authors: Mirabela
authors: M
authors: Hali
authors: Judith A
authors: JA
authors: Whittum-Hudson
authors: Rangaramanujam M
authors: RM
authors: Kannan
authors: Alan P
authors: AP
authors: Hudson
mesh_terms: D002460:Cell Line; D016993:Chlamydophila pneumoniae / Q000378:metabolism*; D002876:Chromosomes, Bacterial / Q000378:metabolism; D004247:DNA / Q000378:metabolism*; D004261:DNA Replication; D050091:Dendrimers / Q000737:chemistry*; D018014:Gene Transfer Techniques; D005798:Genes, Bacterial / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000378:metabolism; D006801:Humans; D018625:Microscopy, Atomic Force; D016366:Open Reading Frames / Q000235:genetics; D010316:Particle Size; D010957:Plasmids; D055672:Static Electricity; D014170:Transformation, Genetic
chemical_list: D050091:Dendrimers; D049452:Green Fluorescent Proteins; D004247:DNA
keywords: Chlamydia; Genetics; Pathogenesis; Transcription; Transformation