Difference between revisions of "UW-Stout/Ultraviolet Light"
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* [https://en.wikipedia.org/wiki/YEPD YPD media], both broth and 2% agar plates] | * [https://en.wikipedia.org/wiki/YEPD YPD media], both broth and 2% agar plates] | ||
* Glycerol stocks of [[UW-Stout/Knockout_Protocol|knockout strains]] | * Glycerol stocks of [[UW-Stout/Knockout_Protocol|knockout strains]] | ||
− | * [https://ecatalog.corning.com/life-sciences/b2c/US/en/Surfaces/Advanced-Cell-Culture-Surfaces/Corning%C2%AE-Treated-Culture-Dishes/p/430166 Cell 60 mm Culture Dish] | + | * [https://ecatalog.corning.com/life-sciences/b2c/US/en/Surfaces/Advanced-Cell-Culture-Surfaces/Corning%C2%AE-Treated-Culture-Dishes/p/430166 Cell 60 mm Culture Dish, containing 6 ml of agar] |
==Equipment== | ==Equipment== | ||
* Incubator set to 30°C. | * Incubator set to 30°C. | ||
− | *Bachur & Associates Sanata Clara, CA 95050 Model LS-100-3 UV Light Exposeure System | + | *[http://www.bachur-n-associates.com/UV%20Exposure%20Systems%20with%20Enclosures%20-%20Nov%202017.html Bachur & Associates Sanata Clara, CA 95050 Model LS-100-3 UV Light Exposeure System] |
==Protocol== | ==Protocol== | ||
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# One day before the experiment, pick a colony from each strain into 5 ml YPD broth in a disposable test-tube. Incubate on the roller drum at 30°C overnight. | # One day before the experiment, pick a colony from each strain into 5 ml YPD broth in a disposable test-tube. Incubate on the roller drum at 30°C overnight. | ||
# The morning of the experiment, dilute each culture to an OD600 of 0.2 in YPD broth. Return to the drum roller for between 30 and 90 minutes. | # The morning of the experiment, dilute each culture to an OD600 of 0.2 in YPD broth. Return to the drum roller for between 30 and 90 minutes. | ||
− | # Transfer 33 µl to a well in the | + | # Transfer 33 µl to a well in the Culture Dish. |
− | # Set up the UV light | + | # Set up the UV light Exposure System as follows: |
#*400 watts | #*400 watts | ||
#*time increments | #*time increments | ||
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==Result== | ==Result== | ||
+ | [[File:UV Control.jpeg|300 px]] | ||
+ | [[File:UV 5 seconds.jpeg|300 px]] | ||
+ | [[File:UV 10 and 15 Seconds.jpeg|300 px]] | ||
+ | [[File:UV 20 and 50 Seconds.jpeg|300 px]] | ||
+ | [[File:UV 60 and 100 Seconds.jpeg|300 px]] | ||
+ | [[File:UV 500 and 900 Seconds.jpeg|300 px]] | ||
+ | [[File:UV 700 seconds and 900 seconds.jpeg|300 px]] | ||
+ | |||
+ | |||
+ | We chose to use the control of 700 seconds given the fact that it had cell death but a small amount of colonies present. |
Latest revision as of 07:05, 2 May 2019
Contents
Materials
- YPD media, both broth and 2% agar plates]
- Glycerol stocks of knockout strains
- Cell 60 mm Culture Dish, containing 6 ml of agar
Equipment
- Incubator set to 30°C.
- Bachur & Associates Sanata Clara, CA 95050 Model LS-100-3 UV Light Exposeure System
Protocol
- Three days before the experiment, streak knockout strains onto YPD plates.
- One day before the experiment, pick a colony from each strain into 5 ml YPD broth in a disposable test-tube. Incubate on the roller drum at 30°C overnight.
- The morning of the experiment, dilute each culture to an OD600 of 0.2 in YPD broth. Return to the drum roller for between 30 and 90 minutes.
- Transfer 33 µl to a well in the Culture Dish.
- Set up the UV light Exposure System as follows:
- 400 watts
- time increments
- place in the incubator in dark for 48 hours
Result
We chose to use the control of 700 seconds given the fact that it had cell death but a small amount of colonies present.