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Growth Of Yeast Biofilms
By Raykia Koroma
Background Information
There is a species of yeast cell called Saccharomyces
cerevisiae that is commonly known as Baker’s Yeast.
In the lab I am working with Saccharomyces
cerevisiae has been genetically engineered to express
the ALS5 gene from another yeast called Candida
albicans.
Candida albicans is a fungus that causes yeast
infections that infect 10 out 1000 people a year. It also
kills hundreds of AIDS victims and cancer victims a
year. It only attacks people with suppressed immune
systems which means it is an opportunistic pathogen.
Candida albicans expresses 6,000 genes one of them is
ALS5. ALS genes encode an adhesion glycoprotein.
There are 8 different ALS genes: ALS1-ALS7 and
ALS9.
Purpose
•I want to know if I express this ALS5 protein
would I get a different or better type of biofilm.
• If yes what does it mean?
•Is ALS5 important for the growth of the biofilm?
Materials
•Shaker
•Plates
•Pipette
•Flask
•Gloves
•Incubator
•Csm-URA liquid
•Autoclave
•Csm- complete liquid
•YPA agar media
•Csm-URA agar media
•Csm-Complete agar media
•W3031B
•ALS5
•ALS1
•ALS3
•PADH
Strains
•Spectrophotometer
Methodology
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I made 500ml of YPA, Csm- URA and Csm- Complete agar media’s.
Split each media into 4 different flasks that contain 125ml in each with
- 2% agar w/galactose
- .3% agar w/galactose
- 2% agar w/glucose
- .3% agar w/glucose
Autoclaved them for 25 minutes.
Poured them on plates and left them overnight in a 30 degrees C
incubator.
Streak
-ALS5 on Csm-trp
-W3031B on YPED
-ALS1 on Csm-URA
-ALS3 on Csm –URA
-PADH on Csm-URA
Inoculated the strains into flasks and put them in a 30 degree C shaker
Measure their Optical Density(OD)
If they all reach .5 great but if not then I would have to spin them down for about
10 minutes.
Dilute so that it’s 3.3 x10 6 cells/ml2 using the Dimensional Analysis technique.
Then inoculate on plates that are divided into either 3 or 2 sections:
-ALS1, ALS3 & PADH same plate with glucose only
-W3031B & ALS5 same plate with glucose or galactose
Leave them to grow at room temperature to grow for about a week
Analysis will be based on the appearance of the biofilm
Results
•No results as of yet.
•I’m expecting for W3031B and PADH to look like
•ALS1 and ALS3 should be a moderate size and be constitutively on .
•ALS5 should grow better on galactose and should look like
Acknowledgements
Dr. Peter Lipke
Janice Lee
Marlyn Gonzales
Raymond Fung
Dr. Sat Bhattacharya
Harlem Children Society Staff
Brooklyn College