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A 7.4-kb plasmid is a common and perhaps essential component of the Chlamydia trachomatis genome. This plasmid occurs as 10 copies per chlamydial chromosomal equivalent. It is unable to replicate in Escherichia coli. Complete plasmid genomes from eight serovars of C. trachomatis have been isolated in E. coli as cloned sequences ligated to pBR322. Restriction enzyme cleavage site mapping indicates that these plasmids are closely related. Homologous plasmid sequences have also been detected by DNA hybridization in all of the 200 clinically isolated strains of C. trachomatis which have been examined. DNA sequences homologous to the C. trachomatis plasmid were not found in eucaryotic DNA nor in a plasmid of similar size isolated from C. psittaci. C. trachomatis plasmid genes are expressed in vivo and the plasmid encoded gene products may play a role in the intracellular growth of this organism. Plasmid encoded genes were also expressed from the cloned C. trachomatis plasmid in E. coli minicells and using an E. coli S-30 in vitro transcription translation extract.