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From 2012.igem.org

I. Inoculate WT B. subtilis Liquid Culture to Test Glycerol Stocks

• Add 5 mL of RT Nutrient Broth to a clean 50 mL conical tube
• Remove 1 WT B. subtilis glycerol stock from the -80 ^oC freezer and put on ice
• Scrape stock ice surface with sterilized inoculating loop
• Inoculate liquid culture
• Return glycerol stock to the -80 ^oC freezer
• Incubate culture at 37 ^oC with shaking overnight with cap loosened