UW-Stout/Heat Shock FA21

Materials Thermocycler 96 channel well Wild-type yeast cells PCR test tubes P20 and P200 micro pipetters Multichannel pipette Media and Plates (all in 475ml H2O) 500 ul of YPD 10g Peptide 5g Yeast Extract 10g Agaar UW-Stout/Heat Shock SP21 - SGD-Wiki (yeastgenome.org) Procedure Media Creation Procedure Measure out 10g of Peptide and put into 500ml jar. Measure out 5g of Yeast Extract and put into 500ml jar. Measure out 10g of Agaar and put into 500ml jar. Then add 475ml of H2O into the 500ml jar. Afterwards bring the jar to a labratory steam sterilizer and put it in for 30 minutes. finally pour the media into plates up to half way. Heatshock Procedure Calibration Experiment Procedure Set thermocycler to 48°C sitting temperature. Vortex the yeast culture briefly to resuspend the yeast cells. Label 6 pcr tubes A through F. Micropipette 90 ul of water into the pcr tubes. Micropipette 10 ul of each strain Label each tube “A, B, C, D, E, F”. Insert all 6 of the tubes into the thermocycler for exactly 42 minutes at 48°C. Afterwards remove all of the tubes from the thermocycler. Dilution Procedure

In a 96 channel well put in your six yeast strains then put them into the left most column desending from A to F.

Then in the rest of the rows add 90 ul of H2O into A-F. After adding the water take 10 ul of the strains add them to the next column and mix. Plating Procedure Label each petri dish accordingly, first plate "A,B,C" and the second plate "D,E,F". Micropipette 5 ul of the strains onto the coordinated petri dish using the . Place all the petri dishes in the incubator (30°C). Let sit for 48 hours. Take out and analyze the growth. Repeat for 3 trials.