Controlled Turgidity; a simple and efficient method for transformation of DNA plasmids into Halobacterium salinarum R1.

Halobacterium salinarum is a model organism used for archaeal genomics studies in many investigations. This well-known microorganism loves hypersaline habitats. However, conventional methods of transformation of DNA for genetic manipulation of the halophile, like electroporation, are impossible, and the current procedure is time-consuming, sensitive, and difficult. Therefore, the development of easy and efficient methods for genetic manipulation, especially the transformation of this species, are important. In this study, we introduced a highly efficient, simple method for H. salinarum transformation. This method is based on the controlled short-time swelling-deswelling of this halobacterium. The halophile was exposed to hypo-osmotic shock by adding deionized water containing the desired plasmid to the cell suspension, which decreased the NaCl concentration to around 2.5M. After 60 seconds, the shock was removed by re- increasing the salt concentration of the suspension to 3.3M. During the osmotic shock, plasmids from the low salt concentrated fluid are sucked through the cell membrane. The transformation was confirmed by PCR and colony count. While the conventional PEG-mediated method is time-consuming and laborious due to the probability of DNA precipitation and cell aggregation by PEG, the present method eliminated PEG and its possible unwanted consequences. With the same amount of initial plasmid, the transformation frequency of the osmotic shock was more abundant in the proposed method compared with the PEG-mediated method. A growth of about 105 transformants/μg of DNA revealed osmotic shock is an efficient transformation method in H. salinarum. This fast and easy transformation method can be used to transform this microorganism and probably other halophiles instead of other current laborious methods.