This page is dedicated to the test protocol used to view stress on yeast cells in a lowered pH environment. We started by mixing up 1 molar HCL into distilled water using a pH probe to determine when we reached the desired level. We had pH from 7 down to 4 going in roughly 1 pH step per solution. We the used these in a calibration test where we tested a unmodified wild type BY4735. This gave us a data sheet showing the growth times and effects of the varied pH levels.
Yeast Cells To test
8 Test Tubes
10 ml of 1 molar ACL
Steps: 1) Gather the test tubes and start to label each one with the pH levels accordingly by increments of 1. And label each tube with the first one at 4 and then 5 and then 6 and then for the control and labels it’s neutral base of 7 and have that be the control of the experiment.
2) Gather the professor and ask him to hover/ help you with the dilution of the sulfuric acid at the according pH levels marked on the test tubes and begin to dilute the acid to the first pH of 4 and then keep diluting the acid by increments of 1 diluted pH to the according tube of labeled up to the ph of 6.
3) Now Either invert the tubes to thoroughly mix the solution or put it on the vortex to make sure it is mixed.
4) Grab the plate and start to pipet out 90 uL of the yeast cell into 12 wells of the plate you are using. And then label Three wells accordingly with the levels of the ph 4 and then 5 and then 6 and then for the base/neutral concentration of 7 just put the cell into the wells without any acid. And then pipet out 10 ul of the first acid concentration of 4 into the labeled well of ph 4 and then same on for the other labeled levels of ph.
5) After it has sit for about a day look at each well and observe the level of growth compared to the control and write down in your notebook on what you see and the level of damage the different levels of ph caused.
Raw Calibration Data:
start of data? When looking at this mess of a graph we must find the useful bits, which to us is going to be the doubling times for each line. In order to find doubling time we must use a doubling time equation which is as follows: Td=(Tf-Ti)*(((log(2))/(log((q2)/(q1))) I will be labeling the data as follows
Time q value Your Favorite Gene: 1: Ex) Ex) 2: Ex) Ex)