The Easiest Yeast Transformation Protocol on Earth

There are several yeast transformation protocols around, and most of them require a lot of steps: overnight starter culture, dilution and growth to logarithmic phase, several washes, and so on…

These protocols work very well since they have been optimised for maximum transformation efficiency, which is needed for applications like library construction.

But they are overkill when you need to do something that doesn’t need such high efficiency, like transforming a standard shuttle vector-based plasmid.

In those situations, a quick, no-fuss (albeit lower efficiency) protocol is what you need. And lucky you, here it is:

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PROTOCOL

Chemically Competent Cells Protocol

Imagine having tube after tube of competent cells for your transformations, all (virtually) free of charge. The way to reach this nirvana is to make your own competent cells. Traditionally this can be a tricky feat, but the procedure laid out in this guide makes it simple and routine.

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1. Choose a large single colony from a plate.

2. Streak on an YPD plate using a singe toothpick and incubate from overnight to up to 48 hours at 30°C. (Note: If your yeast are too old you’ll will see little or no growth – you’ll need to go back to the glycerol stock!).

3. Scrape all freshly grown cells from plate (do not use cells from the beginning of the streak: there is a lot of old, dead meat there). Resuspend in 1 ml of your usual yeast transformation buffer containing lithium acetate.

4. Spin down, discard the supernatant and resuspend in the rest of the buffer.

5. Add your plasmid and single stranded DNA, mix and leave the mixture on your bench for five minutes.

6. Add PEG, invert several times and place in a static incubator at 30°C for an hour.

7. Resuspend by inverting the tube several times, then place at 42°C for 10 minutes.

8. Resuspend again and plate 0.2 ml on a selective plate.

9. Incubate for several days at 30°C.

10. Find something to do with the tons of time you’ve saved!

For a few tips on what can go wrong with a yeast transformation, check out this article. What are your tips for speeding up yeast transformation?

Vicki Doronina

Vicki has a PhD in Molecular biology from the University of Edinburgh.

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