FastYeastTransformation

Protocol: Fast yeast transformation

1. Add 50 μl carrier DNA to a 1.5 ml tube.

2. scrap cells from plate and add to the carrier DNA.

3. Add in the following order:

1. 1-2 μl DNA-plasmid (up to 30-50 μl)

2. 240 μl PEG (50%)

3. 36 μl Li-Ac (1M)

4. resuspend with blue tip

5. incubate at 30°C or RT at least 30 min

6. heat shock: 45°C, 15 min (or 42°C for 20 min)

7. spin down, resuspend in 100μl H₂O and plate everything on corresponding medium

Material:

• Plasmid: 100 ng - 1 μg

• carrier DNA: salmon/herring sperm DNA in H2O, 2mg/ml stock, 500 μl aliquots, heat-inactivated at 95°C for 5-6 min and put on ice before use (has to be done once).

• 50% poly-ethylen-glycol (PEG) (MW 3''350) solution, filter-sterilized

• 1M Li-Acetate, autoclaved