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Transforming Yeast

  1. Grow up some fresh yeast on Y(E)PD media. Using an inoculating loop gentle scrape some off of the plate and very gentle spread it around a fresh plate. Let the plate grow at 30C or less until you see white growth(24-48 hours). The fresh plate should be good for transformations for 1-2 weeks if properly stored at 4C.

 

  1. Scrape yeast off of plate using inoculation loop(enough to fill the loop) and mix into 100uL of the transformation buffer in a micro tube. Transformation buffer: 100mM LiAc, 40% PEG, 10mM Tris, 1mM EDTA, 0.1mg/mL of single stranded salmon sperm DNA

 

  1. Add 0.1 microgram - 1 microgram of plasmid

 

  1. Incubate the micro tube at ~42C for 45 minutes. This can be approximated by letting the tube float in water that is warm but not hot from the faucet.  

 

  1. Incubate the tube at 30C or less for at least 30 minutes and then plate selective media

 

  1. Incubate the plate at 30C or less(room temperature is ok) for at least 2 days or until yeast begin to grow