Sugar DaddYeast
Or sensing sugar with Yeast
In this project, we want to use Saccharomyces cerevisiae (also known as baker’s yeast) as a sugar sensor. We can observe its reaction in two ways, that are induced by different sugar concentrations in their media where it develops.
First because they “eat” sugar in their environment, to do cellular respiration or alcoholic fermentation. To check out these two types of metabolism, just look at this quick video here and here (you’ll even know how to do your own cider!). Basically, they will produce more waste of these metabolisms if more sugar is available to “eat” (just like us humans, if you understand what I mean…).
Second, because they react to higher concentrations of sugar in their environment in a specific way. Indeed, to survive “hyperosmotic pressure”, the phenomenon induced by higher concentrations of a solute in the surroundings of the organism than in the organism itself, they react by producing certain molecules, such as glycerol. If you did not understand well this phenomenon, watch this video or this one- it’s quite long but you’ll know a lot of interesting stuff (that’s the core of our variable, so yeah, that’s important to understand if you want to redo the project !).
At first we wanted to measure the quantity of alcohol (the output of alcoholic fermentation) produced, but we decided that it was way too difficult to measure, as well as CO2, the output of cellular respiration.
So we searched a way to spot the reaction of the cells to hyperosmotic pressure.
And we found a yeast’s protein, called HOG1. Proteins are the basis of every living form. They are coded by genes. Every gene codes for one protein. Now that’s very basic, if you want to know more about proteins and genes, you should think about following a molecular biology course. Or check out this video for an introduction.
This protein relocates from the cytoplasm into the nucleus of the yeast when the yeast is under hyperosmotic pressure.
We used a Green Fluorescent Protein (GFP) modified strain of yeast to spot the relocation of this protein, HOG1.
GFP-modified strains of yeast are GMOs. It means that their genetic information has been changed. Basically, bio-engineers will insert a gene coding for a fluorescent (which means, basically, that it glows back when exposed to a certain light) protein just next to a gene of interest -here, the gene coding for HOG1. We can observe a specific protein glowing with a fluorescence microscope, at a specific location of the cell.
Our protocol was simple : we would mix 10µL of yeast media (YPD) with a certain concentration of glucose with 10µL of an overnight culture of a GFP-HOG1 yeast strain. Then, we would put 3µL of the mix on a microscope slide, and observe this mix at the fluorescence microscope in less than 5 minutes. We would take several pictures (10-20), and we would count the number of cells, for each slide, that have fluorescence clearly and obviously more in the nucleus than in the cytoplasm, and the other.
Then, from this counting of cells, we would get a percentage. Here are our results, simplified for you to understand faster :
What can we conclude about that ? Not much. First, we don’t have enough data to be statistically relevant. Moreover, we have many experimental errors in our protocol. Did you find them all ?
First, we have a subjective classification of the cells that fluoresce in the nucleus and ones that fluoresce in the cytoplasm.
With a computer script to automatically treat the picture, we would have fewer variations in our measures.
Even though we have lots of experimental errors and confirmation biases (to know what it is, go there), we can observe from our results that hyperosmotic pressure does have an impact on the relocation of HOG1 in the nucleus, from 10% of glucose in the media. It’s also the concentration with the more response of the yeast cells. We observe a decrease of the response in higher concentrations, which may be due to the harsh conditions of such osmotic pressures that would kill the cells.
Further experiments should be performed, with less errors and biases, and more repetitions. The goal would be to understand if a HOG1-GFP strain of Saccharomyces cerevisiae can be used as a really accurate, precise and with a good resolution hyperosmotic pressure sensor!
Thanks a lot for reading our blog post, we hope you understood it. Don’t hesitate to contact us if you have any question, by email or twitter!
@leonfaurefdv
@louise_jacquot
@NinaVarcha
Link to the GitHub documentation : https://github.com/
Image sources : fr.wikimedia.org
No comments:
Post a Comment