Transformation of Xylella fastidiosa using replicative shuttle vector pUFR047

• Author(s): Gabriel, Dean; Harakava, R.; Vanamala, A.;
• Abstract: Xylella fastidiosa (Xf) is a xylem-inhabiting bacterium that causes serious diseases in a wide range of plant species. Two of the most serious of these are Pierces Disease (PD) of grape and Citrus Variegated Chlorosis (CVC). Both PD and CVC genomes have been completely sequenced. Functional genomic analyses of Xf have been severely limited by lack of a stable replicative shuttle vector, pUFR047 is a small, stable, wide host range, conjugative, replicative, repW shuttle vector with a long history of use in Xanthomonas. pUFR047 was transferred into a rifampicin-resistant Florida PD strain, PD1A, by both conjugation and electroporation, pUFR047 was also electroporated into the sequenced California PD strain Temecula. Transfer by conjugation was inefficient due to background growth of E. coli. Transfer by electroporation occurred with or without DNA inserts into both PD strains at a frequency of ca. 50 transformants/microgram DNA. The vector was replicative in both Xf strains and was reisolated after ten generations of growth and used to transform E. coli. Stability tests are underway.
• Publication Date: Jun 2002
• Journal: Phytopathology