Efficient DNA transformation of bacterial competent cells has a critical role in biotechnology experiments such as gene cloning and protein expression. Transformation rates are commonly associated with the competency state of the bacterial cells. In this paper, it was demonstrated that transformation efficiency is strongly related to the growth phase of bacterial culture used for competent cell preparation, as well as special chemical treatments applied to make them competent. Results showed that exposure of the cells from the late growth phase (OD600; 0.9-0.94) to monovalent and divalent cations resulted in the highest transformation efficiencies (about 100-fold). These competent cells are particularly useful for the transformation of ligated and large DNA molecules (more than 10 kbp).