McClean: Cycloheximide Stock Solution
This is the standard 50mg/ml stock solution. This is diluted to 100μg/mL to inhibit translation (though you should check this concentration does what you think it does whenever you develop a new protocol using cycloheximide). Each person in the lab makes up their own stock of cycloheximide and keeps it in their own -20°C box.
- Cycloheximide (MP Biomedicals, Cat #100183)
- DMSO (Fluka 41639, Ultra for molecular biology; stored in the Flammables cabinet)
Weigh out the appropriate amount of cycloheximide and add it to the appropriate amount of DMSO to get a 50mg/ml solution. Use the Botstein lab's balance as it is much more accurate than ours. It seems to make the most sense to aim for about 100mg (but if it is slightly over or under that is fine) and then put this in a 2ml eppendorf or similar tube (you can use a 15ml conical if you are going to make a larger volume) and add the appropriate amount of DMSO to this based on what you weighed out.
Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!
*Megan N McClean: I generally find it useful to make a series of 1:10 dilutions of the α-factor (1mg/mL, 100μg/mL, 10μg/mL, 1μg/mL) when I first make the stock, as these are 1000x the concentrations I generally start out with for most experiments (so that I can add 1μL of stock solution per ml of media). I usually make 900μL of all of these dilute stocks (ie, start with 100μL of 1mg/ml stock +900μL of DMSO for the 100μg/mL stock, from that take 100μL and add it to another 900μL of DMSO for the 10μg/mL, and then once more for the 1μg/mL stock). I then divide the 900μL into 100μL aliquots so that I don't have to freeze/thaw the entire stock too often.
Please sign your name to your note by adding '''*~~~~''': to the beginning of your tip.
- Megan N McClean 14:01, 6 October 2011 (EDT)
or instead, discuss this protocol.