Versatile Methodology for Efficient Large-sized DNA Delivery Between Microorganisms Without In vitro Purification

Purified DNA plasmids traditionally used for microbial transformation have been supplanted by extracellular plasmids released via host bacterial lysis, offering an alternative approach for DNA-plasmid delivery. Specifically, shuttle vector plasmids liberated from host Bacillus subtilis were directly employed for the transformation of chemically competent cells Escherichia coli, eliminating the need for biochemical purification. This unconventional DNA delivery technique, referred to as ’Cell Lysis Technology to provide Transformable Extra-cellular DNA; CELyTED’, has been successfully adapted for the transformation of microorganism Saccharomyces cerevisiae as well. The protocol includes optimized conditions for efficient cell lysis of the donor host cells. Notably, ’ CELyTED ’ enables the introduction of large-sized DNA plasmids exceeding 50 kb into target microorganisms mitigating the potential adverse effects of physical shearing during the purification process. This simplicity in the delivery protocol makes it versatile for both prokaryotic and eukaryotic microorganisms, establishing a fundamental platform in the synthetic genome field. Our study demonstrates the feasibility of introducing large DNA plasmids into cells E. coli and S. cerevisiae using the lysate of donor host cells, showcasing the potential of ‘CELyTED ’ as a streamlined approach in genetic transformation methodologies.