McClean: Sonication Yeast
Protocol for sonicating yeast for microscopy (and other applications) where you want the sonication to maximally break up clumps while maintaining 100% viability.
Our sonicator is a Fisher Scientific Model 705 Sonic Dismembrator.
- Mid-log (or other stage of growth) cell culture
- 1.5 ml eppendorfs
- Hearing protection
- Clean the sonicator microtip by wiping it down with 70% ethanol and a kimwipe
- Aliquot ~500 μL cells into 1.5 ml eppendorf tube. DO NOT USE A GLASS TUBE
- Submerge the sonicator microtip into the eppendorf.
- Aim to have the tip ~ 1 cm from the bottom of the tube. Too deep and you won't get adequate mixing. Too shallow and you run the risk of foaming
- Center the tip inside the eppendorf
- MAKE SURE THE TIP IS NOT TOUCHING THE WALLS OR BOTTOM OF THE TUBE
- Turn on the sonicator
- Press "Yes" that you are using a microtip
- Select to modify a program or sequence
- Select/Modify a program
- Program 1 has the Maitreya lab protocol saved
- Amplitude: 5
- Pulse-ON Time: 1 sec
- Pulse-OFF Time: 1 sec
- Elapsed Time: 20 sec
- WEAR EAR PROTECTION
- Press Start
- Monitor the tube to make sure there is no foaming. If foaming does occur:
- Stop the sonicator
- Centrifuge the tube until the foam has dissipated
- Vortex to resuspend the cells
- After sonication, clean the microtip with 70% ethanol and kimwipe
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.
*Megan N McClean 18:52, 8 March 2017 (EST) Two places to start in determining a good protocol for our QSonica sonication protocol are below. The Botstein Protocol is the one that I used previously at Princeton. Maitreya's seems very similar except for longer pulses and longer rests.
Botstein Lab Protocol: Sonicator unknown, Program #1, 5 sec pulse, pulse on 0.5sec, off 0.5 sec, output level 5
Maitreya Lab protocol: Misonix S4000, Program #1, which consists of 10, 1 second bursts at Amplitude=5, with a 1 second rest in between bursts. This seems to separate cells nicely
- Taylor D. Scott 16:56, 20 March 2017 (EDT): I tested the Maitreya lab protocol (amplitude = 5, 1 sec on, 1 sec off, 10 periods) on a Sigma strain and it separated the cells nicely. I had 4% dead cells after sonication.
- Megan N McClean 14:01, 20 July 2011 (EDT)
or instead, discuss this protocol.