<p> Freezer Storage of Bacterial Strains</p><p>1. Grow a 2 ml culture of the strain to be stored in LB + appropriate antibiotic. </p><p>2. In a 2.0 ml NALGENE CRYOVIAL, combine 1ml overnight culture 0.5 ml Medium E + 21% DMSO.</p><p>3. Mix with pipettor and store at -75°C.</p><p>4. When subculturing from freezer stocks, it is not necessary to thaw the tube containing the bacteria. Just touch the loop to the surface of the frozen culture. Never inoculate a liquid culture directly from a freezer stock. This may lead to loss of the plasmid, and or contamination of the liquid culture.</p><p>50 x Medium E (800ml)</p><p>MgSO 4 • 7H2O 8 g Citric Acid • H2O 80 g K2HPO4 (Anhydrous) 400 g NaNH4HPO4 • H2O 140 g Add H2O to 800 ml. Autoclave</p><p>1x Medium E + 21% DMSO</p><p>21 ml DMSO 79 ml 1x Medium E Filter Sterilize</p>