Herskowitz Lab Protocol
From : RKT (7/96)

Yeast Transformation

  1. Grow 100ml cells to A600 = 0.5-0.7 (A600 0.7 = 2 X 107 cells/ml)
  2. Pellet 5' 3K
  3. Wash w/ 5ml LiAc mix;
  4. Pellet
  5. Resuspend in 1ml LiAc mix
  6. Transform : Use 100ul cells (in eppendorf)/ tformation. Add 1ug DNA in <15ul and 50ug carrier DNA
  7. PEG : Add 0.7 ml PEG mix
  8. Mix by vortexing
  9. Incubate 30' at 30°C
  10. Heat Shock 15' -20' at 42°C
  11. Pellet 5" in microfuge (LM-pellet 30")
  12. Resuspend using yellow pipette tip to disperse pellet in 200ul water
  13. Plate

Materials

LiAc mix : 100mM LiAc in TE (1.02g/100ml)

PEG mix : 40% PEG in 100mM LiAC/TE = 4g of PEG in 7ml - made fresh

carrier DNA : Calf thymus or salmon sperm DNA that has been autoclaved, boil fresh before use for approxiamately 4 minutes

Other online versions available from the Botstein Lab, Gottschling Lab.

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