Team:BYU Provo/Notebook/Phage Purification/Winterexp/Period4/Dailylog

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| colspan="3" | <font color="#333399" size="5" font face="Calibri"> '''Phage Purification March - April Notebook: April 1 - April 14 Daily Log'''</font>
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| colspan="3" | <font color="#333399" size="5" font face="Calibri"> '''Phage Purification March - April Notebook: April 15 - April 30 Daily Log'''</font>
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: [[Team:BYU_Provo/Phage_Purification|Overview]]
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: <u> '''Phage Purification''' </u> </font>
: [[Team:BYU Provo/Notebook/Phage_Purification/Winterexp|March-April]]
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<font size="4"> '''4/15/13''' </font>
<font size="4"> '''4/15/13''' </font>
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:I feel like we could have been a bit more engaging and better rehearsed as well.  
:I feel like we could have been a bit more engaging and better rehearsed as well.  
:We will need to find a way to test the capsids for viability, and somehow characterize them.
:We will need to find a way to test the capsids for viability, and somehow characterize them.
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<font size="4"> '''4/2/13''' </font>
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<font size="4"> '''04/16/13 - 04/30/13''' </font>
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-Liquid culture 4/2/2013
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We put a colony from our W3110 plate of E.coli into 1mL LB broth and placed in the 37 C incubator overnight. We did this with 5 colonies.
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-Results: the control tube of broth showed no contamination.
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-We also put 500microL of 40% glycerol into cryovials for use tomorrow to prepare freezer sotcks.
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- Spring Break
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<font size="4"> '''4/3/13''' </font>
 
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- Today we will be setting up an overnight culture of E. coli, to grow for use with phage amplification.
 
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:We will take a 500 flask; with 50 mL LB broth, and place a culture from the W3310 E.coli culture plate to grow overnight in the 37 C incubator
 
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:We will create a glycerol stock of the E.coli culture to save in the -80 C freezer.
 
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<font size="4"> '''4/4/13''' </font>
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- We made glycerol stocks W3110 E. Coli
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:We put .5mL glycerol in cryovials and .5mL liquid culture from the overnights prepared yesterday.
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:We vortexed each tube for 10 seconds and then we stored the tubes in the -80°C freezer in Dr. Grose’s lab.
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<font size="4"> '''4/5/13''' </font>
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-We did spot tests of phage on E. Coli BL21
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:We plated together .5mL of BL21 with 1XLBTA.
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:We separated the plates into different segments and then spot tested the following phages.
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::14 T3
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::13 T5
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::10 T2
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::20OX171
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::40ØX174
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:The plates were then put in a 37°C incubator
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-Results:
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:There were no plaques on any of the plates.
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:There was no contamination in the control quadrant.
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:A uniform bacterial lawn formed with no phage infection.
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<font size="4"> '''4/8/13''' </font>
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-We performed a phage titer test to determine which bacteria would be most viable for our phage propagation.  
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:We used BL21 and W3110 strains of E. Coli.
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:We placed 100 µL of broth into 5 ependorf tubes.
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:We used as phage 10 µL of 1L, 10L, 40T4, T4DOS, and T4 infected phage and placed them each in one of tubes labeled -1.
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:We performed a dilution series taking out 10 µL each time and placing it into the next tube, 5 times. The total volume in the last tube was 110 µL.
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:We added 4.5 mL of 1X top agar to .5 mL of broth and plated it on LB plates.
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:We spotted each concentration on the plates and incubated it overnight at 37°C.
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-Results from 04/08:
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:Phage 10L w/ w3110 had large scale infection every concentration
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:Phage 10L w/ BL21 had infection in very large concentrations
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:Phage 1L w/ w3110 had infection in very large concentrations
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:Phage 1L w/ BL21 had infection in very large concentrations
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:Phage T4 DOS w/ w3110 had infection in very large concentrations
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:Phage T4 DOS w/ BL21 had infection in very large concentrations
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:Phage 40 T4 w/ w311o had no phage infection
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:Phage 40 T4 w/ BL21 had large scale infection at every concentration
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:Phage T4 inf w/ w311o had large scale infection at every concentration
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:Phage T4 inf w/ BL21 had large scale infection at every concentration
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:We had a significant amount of running that occurred on almost every plate, so the results were a little difficult to read. We concluded that the phage is a high enough concentration that we would have to do another phage titer to a smaller dilution in order to determine actual concentration. The controls were all a lawn of bacteria.
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<font size="4"> '''4/10/13''' </font>
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-We performed titers of T4 infected phage from off of two separate E. Coli strands, BL21 and W3110.
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:We added .1mL of the liquid culture to 1 mL of both strands of bacteria.
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:After allowing infection to occur overnight, we centrifuged the tubes to separate the bacteria from the phage.
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:We separated the supernatant into a new eppendorf tube, and used 10 µL as the 0 concentration.
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:We then performed a phage titer using five tubes of 90 µL LB each, adding 10 µL from each one to the next.
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:Using a micropipette we spotted 2µL of each dilution onto both 4.5 ml 1x plates and .5 mL W3110 plates.
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:To avoid smearing, we allowed the plates to sit for over an hour and then we incubated them at 37 C overnight.
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-Results:
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:The concentrations we used are still way too strong. We will have to dilute the concentration even further if we want to see anything more than cleared plaques.
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<font size="4"> '''4/12/13''' </font>
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-We watched presentations from the Cholera Group Today
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Latest revision as of 00:02, 28 September 2013


Phage Purification March - April Notebook: April 15 - April 30 Daily Log



Phage Purification
March-April
May-June
July-August
September-October



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4/15/13

-We presented our project today. -There were a few things that we needed to do better.

For one, we need to present better background and show how it ties into our research, specifically the need for a capsid library.
I feel like we could have been a bit more engaging and better rehearsed as well.
We will need to find a way to test the capsids for viability, and somehow characterize them.


04/16/13 - 04/30/13

- Spring Break

<< Previous Next >>

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