I am studying a clinical Staphylococcus aureus isolate which has special phenotype. In a further study, I plan to manipulate genes like knockout or over-expression in the isolate. However, I tried to transform even 20 ug plasmids into the isolate several times but failed. I tried both shuttle plasmid extracted from S. aureus RN4220 and Staphylococcus plasmid like pT181. The isolate has no plasmid.
Any suggestion or tools for transformation of clinical Staphylococcus aureus isolates?
Maybe you can try this...Simple DNA transformation in Pseudomonas based on the Yoshida effect. Rodríguez-Beltrán J, Elabed H, Gaddour K, Blázquez J, Rodríguez-Rojas A. J Microbiol Methods. 2012 May;89(2):95-8. Epub 2012 Mar 3. PMID: 22405834. Pubmed
Usually electroporation can be used. However using clinical isolates it is sometimes difficult to get electro competent cells. An alternative approach might be phage transduction using phages able to transduce between RN4220 and your strain of interest. The strain of interest must not necessarily display plaques with the phage. Sometimes the transduction also works without visible plaque formation. We have done this several times in S. epidermidis and it should work also in S. aureus.
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