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Team:BYU Provo/Notebook/Phage Purification/Fallexp/Period3/Dailylog
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| - | : '''Phage Purification September - October Notebook: | + | : '''Phage Purification September - October Notebook: October 21- 27 Daily Log'''</font> |
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| + | : <u> '''Phage Purification''' </u> </font> | ||
: [[Team:BYU Provo/Notebook/Phage_Purification/Winterexp|March-April]] | : [[Team:BYU Provo/Notebook/Phage_Purification/Winterexp|March-April]] | ||
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| - | <font size="4"> ''' | + | <font size="4"> '''10/21/13''' </font> |
| - | - | + | -Today we ran PCR on T4 wild type and mutant phage to determine the sequence differences in the capsid proteins. |
| - | - | + | :: - [[Team:BYU_Provo/Notebook/Phage_Purification/Fallexp/Period1/Exp/10.21T4PCR|10.21 T4 PCR]] |
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| - | <font size="4"> ''' | + | <font size="4"> '''10/23/13''' </font> |
| - | + | -Ran a gel on the PCR we did last time. | |
| + | : -Gel | ||
| + | :: 100 mL TAE 1x | ||
| + | :: 1 g regular agarose | ||
| + | :: 2 drops ethidium bromide | ||
| + | : Combine and melt TAE and agarose | ||
| + | : Cool and add 2 drops ethidium bromide | ||
| + | : Allow gel to form with 14 well mold (20 minutes in the fridge) | ||
| + | : -Preparing machine | ||
| + | : Fill machine with TAE buffer until it covers gel | ||
| + | : 2 microL DNA ladder in first well | ||
| + | : 2 microL stain and 5 microL sample, mix and put in well | ||
| + | : Run at 130 V for 60 minutes | ||
| - | + | We place the samples in the order: DNA ladder, 297/298 W M, 301/302 W M, 311/312 W M, 313/314 W M | |
| + | W=wildtype M=mutant | ||
| - | + | We got bands for both the 311/312 and 313/314 wild type and mutant. We need to do PCR cleanup and start with the cloning procedure. 297/298 and 301/302 didn't show any bands for either the wild type or the mutant. We suspect this may be to boiling them for too long (12 min) at the start. We are going to try boiling them for only 5 minutes and see if we can get bands next time. | |
| - | + | The PCR products were stored in the Cholera QS II box in the freezer. | |
| + | |||
| + | [[File:PCR gel cloning.jpg]] | ||
| + | |||
| + | AB DL AC | ||
<br> | <br> | ||
| - | <font size="4"> ''' | + | <font size="4"> '''10/25/13''' </font> |
| - | + | -Today we ran PCR again on T4 wild type and mutant phage to determine the sequence differences in the capsid proteins. | |
| + | :: - [[Team:BYU_Provo/Notebook/Phage_Purification/Fallexp/Period1/Exp/10.25T4PCR|10.25 T4 PCR]] | ||
| + | |||
| + | AB DL AC | ||
<br> | <br> | ||
| - | <font size="4"> ''' | + | <font size="4"> '''10/28/13''' </font> |
| - | + | - We ran another PCR attempting to get bands for sequencing both the major and minor capsid proteins. We used our normal protocol except did not boil the phage this time. Skip also used the protocol he normally uses to see if reagents are the problem with us not getting a band. | |
| - | + | [[File:T4 gel Oct 28.jpg]] | |
| - | + | - We sent in the major and minor capsid protein genes of the wild type and mutant for sequencing. | |
| - | + | AB DL AC | |
| - | < | + | <br> |
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Latest revision as of 03:22, 29 October 2013
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10/21/13 -Today we ran PCR on T4 wild type and mutant phage to determine the sequence differences in the capsid proteins. AB DL AC
10/23/13 -Ran a gel on the PCR we did last time.
We place the samples in the order: DNA ladder, 297/298 W M, 301/302 W M, 311/312 W M, 313/314 W M W=wildtype M=mutant We got bands for both the 311/312 and 313/314 wild type and mutant. We need to do PCR cleanup and start with the cloning procedure. 297/298 and 301/302 didn't show any bands for either the wild type or the mutant. We suspect this may be to boiling them for too long (12 min) at the start. We are going to try boiling them for only 5 minutes and see if we can get bands next time. The PCR products were stored in the Cholera QS II box in the freezer.
AB DL AC
10/25/13 -Today we ran PCR again on T4 wild type and mutant phage to determine the sequence differences in the capsid proteins. AB DL AC
10/28/13 - We ran another PCR attempting to get bands for sequencing both the major and minor capsid proteins. We used our normal protocol except did not boil the phage this time. Skip also used the protocol he normally uses to see if reagents are the problem with us not getting a band.
- We sent in the major and minor capsid protein genes of the wild type and mutant for sequencing. AB DL AC
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