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Several factors including the culture temperature, bivalent ion, and osmotic stress were gradually optimized for preparing efficient Escherichia coli competent cells. The effect of culture temperature on the transformation efficiency (TrE) of E. coli DH5α was tested with 100 mM CaCl2. The lower culture temperature at 18 °C resulted in higher TrE of 2.5 × 106 cfu/μg, which was about 3.5 times of that...
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