Team:BYU Provo/Notebook/SmallPhage/Springexp/Period2/Exp/5.20 Mutagen Concentration Experiment
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7) Large plaque confirmation (6.10) | 7) Large plaque confirmation (6.10) | ||
- | : From the 45 plates above we picked 12 of the biggest plaques and plated them using x8 top agar to confirm whether the supposed mutant have stable phenotype. Specifically, we added 50uL of LB to 12 eppendorf tubes. Each plaque was then stabbed with a pipet and the pipet was dipped into the 50uL LB; we pipetted up and down several times to ensure that phage particles got in the LB | + | : From the 45 plates above we picked 12 of the biggest plaques and plated them using x8 top agar to confirm whether the supposed mutant have stable phenotype. Specifically, we added 50uL of LB to 12 eppendorf tubes. Each plaque was then stabbed with a pipet and the pipet was dipped into the 50uL LB; we pipetted up and down several times to ensure that phage particles got in the LB. |
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+ | : To 12 test tubes, we added 0.75mL of Bl21 overnight and 30uL of phage in LB prepared in the previous step. They were incubated for 20 minutes. | ||
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+ | : To each of the test tubes, 7mL of x8 top agar was added and then plated unto LB plates. | ||
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+ | : The plates were incubated at 37C for approximately 24 hours. | ||
'''V) Results''' | '''V) Results''' | ||
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: -6 at x6 only showed a couple plaques on each plate. For -4 at x6, there were many plaques. It seems like the number of plaques increase as we increase the mutagen concentration. | : -6 at x6 only showed a couple plaques on each plate. For -4 at x6, there were many plaques. It seems like the number of plaques increase as we increase the mutagen concentration. | ||
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+ | 5) Preliminary selection test - third round | ||
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+ | : x6 top agar produced bigger/more plaques than x8 top agar. For selection purposes, we decided to use x8 top agar from now on. | ||
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+ | 6) Selection - round 1 | ||
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+ | : We saw a few bigger plaques (compared to the average plaque sizes) from the 45 plates. These are ones we are interested in. | ||
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+ | 7) Large plaque confirmation | ||
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+ | : Large scale contamination prevented any plaque from showing up. | ||
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Revision as of 00:25, 13 June 2013
Small Phage May - June Notebook: Experiments
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5.20 Mutagen Concentration Experiment
I) Purpose
II) Expected Outcome
III) Reagents Used
IV) Procedure 1) Applying the mutagen (5.20)
2) Spot test (5.22)
3) Preliminary selection test (5.24)
4) Preliminary selection test - second round (5.29)
5) Preliminary selection test - third round (6.3)
6) Selection - round 1 (6.7)
7) Large plaque confirmation (6.10)
V) Results 1) Applying mutagen
2) Spot test
3) Preliminary selection test
4) Preliminary selection test 2
5) Preliminary selection test - third round
6) Selection - round 1
7) Large plaque confirmation
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