Difference between revisions of "UW-Stout/pH Acid SP22"
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==Introduction== | ==Introduction== | ||
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Our protocol is designed to determine if the knocked-out gene affects or does not affect the growth of our yeasts in environments of pH 7. The pH of 7 was predetermined in a pilot experiment to be a pH that would stress the cell’s growth without killing all the cells. A citric acid monohydrate buffer was used to maintain a pH of 7 in the growth environments throughout the entire growth process. | Our protocol is designed to determine if the knocked-out gene affects or does not affect the growth of our yeasts in environments of pH 7. The pH of 7 was predetermined in a pilot experiment to be a pH that would stress the cell’s growth without killing all the cells. A citric acid monohydrate buffer was used to maintain a pH of 7 in the growth environments throughout the entire growth process. | ||
| − | == Materials/Equipment == | + | == Materials/Equipment== |
• Citric Acid Monohydrate | • Citric Acid Monohydrate | ||
• Disodium Phosphate | • Disodium Phosphate | ||
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| − | + | ==Procedure== | |
'''Preparing Buffer Stock Solutions (0.1M citric acid, 0.2M disodium phosphate)''' | '''Preparing Buffer Stock Solutions (0.1M citric acid, 0.2M disodium phosphate)''' | ||
Revision as of 11:55, 28 April 2022
Introduction
Our protocol is designed to determine if the knocked-out gene affects or does not affect the growth of our yeasts in environments of pH 7. The pH of 7 was predetermined in a pilot experiment to be a pH that would stress the cell’s growth without killing all the cells. A citric acid monohydrate buffer was used to maintain a pH of 7 in the growth environments throughout the entire growth process.
Materials/Equipment
• Citric Acid Monohydrate • Disodium Phosphate • De-ionized Water • Micropipette • 15ml tube (3x) • 96-well clear flat-bottom assay plate • Micropipette tips • Gloves • Incubator set to 30 C • Scale • Sterile cabinet • pH test strips • Yeast strains
Procedure
Preparing Buffer Stock Solutions (0.1M citric acid, 0.2M disodium phosphate) 1. To obtain 0.1M of citric acid monohydrate, combine .21g of in 10ml of de-ionized water 2. To obtain 0.2M of disodium phosphate, combine .28g in 10 ml de-ionized water
Preparing Buffer Solution
1. To create a buffer solution at a specific pH, you must combine x ml of citric acid monohydrate with y ml of disodium phosphate and shake well
pH x ml 0.1M-Citric Acid y ml 0.2M-Na2HPO4
3 79.45 20.55
4 61.45 38.55
5 48.5 51.5
6 38.86 63.15
7 17.65 82.35
== Calibration Experiment protocol ==
== Final Experiment protocol ==
