then we'll compare the the "fitness" of our evolved vs ancestral yeast strains

we're going to be using 8 different strains of yeast.

one is colorless and the other 7 display a color when plated.

I'll maintain the control which will be the colorless strain grown in only  
YEPD (yeast extract peptone dextrose...a medium for yeast growth)
with no acetic acid added.

​you'll be divided into 7 groups.
each group will select a single strain (color) of yeast.  

finally, we'll perform a whole-genome sequencing which will allow us to  identify mutations in our evolved strains 

CO2 + alcohol

yeast are a type of fungi that are capable of a form of anaerobic respiration called fermentation.
Acetic acid is a byproduct of yeast fermentation.

We'll be directing the evolution of acetic acid tolerance in a yeast strain called S. cerevisiae
via artificial selection.

week 3: September 6th
​you will compare your experimental strain to the control for growth and then pass it (P3) into a new (labelled) vial containing either 50mM or 75 mM acetic acid
​in YPD.  The concentration of the HAc you use will be determined based on how your cultures are growing. ​If they look like there's some good growth,
you'll pass to a 75mM HAc tube. 

Bio 111 2023
​CURE

yeast evolution 

via artificial selection

The U.S. produces (2019) 60% percent more milk from 30 percent fewer cows than it produced in 1967.
Each cow produces >2.5 times the milk compared to
cows just a few decades ago.
Via intentional breeding aimed at increasing milk production/profit, Holstein cows now produce about
75 pounds or 9 gallons of milk
every.single.day.
A Holstein calf only needs
9 pounds or 1.25 gallons of milk per day. 

and during our last lab,
we'll do a competition assay...
here's the protocol...

​
​

artificial selection?? for example...

week 4: September 13th
​you will compare your experimental strain to the control for growth and then pass it (P4) into a new (labelled) vial

​ of 75 mM HAc.

      25           50           75             100
​                           mM HAc

week 2: August 30th
​you will compare your experimental strain to the control for growth and then pass it again (P2) into a new (labelled) vial of
​50 mM acetic acid (HAc) in YPD.

yeast growth is normally  arrested above 50 millimolar acetic acid

<-- acetic acid

yeast 

week 1: August 23rd
​you will transfer your yeast into a solution that is 50 millimolar acetic acid.

label your vial:

strain (color)
course section (370)
date
passage number (this week is P1)
​acetic acid (HAc) concentration

glucose

here's the procedure...

 GENTLY vortex the yeast culture you've been assigned until it looks
evenly suspended
(be really careful here - the liquid WILL come out of the top)
 
remove the cap.  do not place it down on the bench

open the sterile swab, avoid touching the cotton tip to anything, and place it into the mixed YPD/yeast mix. swirl the swab around.

remove the cap of the new tube containing 50 mM acetic acid (HAc). 
do not place the cap down on the bench.

move the swab from the yeast culture tube into the new tube containing the 50 mM acetic acid and swirl the swab around gently

remove the swab.
replace the cap as quickly as possible after the swab has been removed.

 you may gently push the cotton tip against the side of the tube to squeeze out any absorbed yeast/liquid
the used swab can then be thrown away

make sure your tube is labelled clearly as previously described.

Place your tube into the 30C incubator 

udders aside,  we're going to explore artificial selection in yeast...

and each week to follow...
​  maybe 100 mM HAc??
What do you think?

Acetic acid accumulation leads to a slow down of growth in yeast. Acetic acid (think vinegar) inhibits the growth of yeast by causing yeast cells to use energy on repair rather than growth and by altering specific chemicals within the cell to signal apoptosis = cell death.