McClean:Making and Using Frozen Yeast Competant Cells
The following is a protocol to freeze down and use any of our S. cerevisae strains for standard LiOAc transformation at a later date. This is especially useful when you often need to perform many transformations using a single background strain. Adapted from the protocol by Geitz et. al 2007, Nature Protocols.
- 50 mL Conical Tubes
- Frozen Competant Cell Solution (recipe to follow)
- Liquid YPD media (From media facility, or recipe in Amberg et. al's Methods in Yeast Genetics)
- 100 well styrofoam freezer racks (that will fit standard 1.5 mL 'minifuge' tubes
- A 250 mL sterile culture flask for each strain
- A 2 L sterile flask for each strain
Frozen Competant Cell (FCC) Solution
Solution is 5% (v/v) glycerol and 10% (v/v) DMSO. Be sure to filter-sterilize.
- Innoculate an overnight culture of your strain into 25 mL of YPD and grow at 30°C and 200 RPM overnight or for 12-16 hours.
Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!
- List troubleshooting tips here.
- You can also link to FAQs/tips provided by other sources such as the manufacturer or other websites.
- Anecdotal observations that might be of use to others can also be posted here.
Please sign your name to your note by adding '''*~~~~''': to the beginning of your tip.
Gietz, R.D. and R.A. Woods. (2002) TRANSFORMATION OF YEAST BY THE Liac/SS CARRIER DNA/PEG METHOD. Methods in Enzymology 350: 87-96.
- Megan N McClean 14:01, 20 July 2011 (EDT)
or instead, discuss this protocol.