Overview
This is the standard 1mg/ml stock solution we use for inducing the pheromone pathway in budding yeast. Generally, we find that the different batches of pheromone can be somewhat variable so it is a good idea to make up a stock that will last you through your experiment and keep it in your own -20°C box. If you have to switch stocks mid-experiment you need to do some calibration experiments to make sure the activity is the same.
Materials
- α1-Mating Factor acetate salt (Sigma T6901-1mg)
- DMSO (Fluka 41639, Ultra for molecular biology; stored in the Flammables cabinet)
Procedure
Shake the new vial of α-factor to get all of the power on the bottom of the vial. Remove the lid and pipet 1ml of DMSO into the vial of α-factor from Sigma. Pipet up and down put the lid back on and swirl to dissolve all of the alpha-factor into the DMSO. Then remove the liquid and aliquot it in 100μL Eppendorf into individual Eppendorf and store these in your own -20°C box. Keep track of the lot # of the pheromone both with your stock solutions and when you do your experiments.
Notes
- Make sure to keep track of lot numbers (write it down in your notebook, on your stock solutions, and each time you do an experiment with α-factor). Really. Please do this!
- 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.