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	===7/10/13===		===7/10/13===
		+	Today we are trying out a different but similar protocol. We are hoping to see clearage in our overnight cultures which we haven't seen so far. This new protocol describes that the phage will super infect the bacteria this would explain why our phage wasn't clearing the flask. The paper states,
		+	:"After one round of infection and lysis,all remaining cells become infected. Some of these
		+	:lyse on schedule, liberating phage which superinfect the majority of cells which are still unlysed. This
		+	:superinfection causes inhibition of lysis (9). Starting at 150 min, samples (5 ml) are removed periodically
		+	:and shaken with chloroform to test for the degree of lysis which can be induced. When that test shows that
		+	:most of the cells can be induced to lyse so that clearing is nearly complete (usually at about 180 min
		+	:after initial infection),the culture is chilled in an ice bath and the pregnant bacteria are pelleted by
		+	:centrifugation at about 5,000 rpm for 10 min in a cold centrifuge."
		+
		+	So today we added 100ul of adenine and 200ul uricil to 25ml flasks of e.coli B in LB and in M9+ minimal media along with 500 ul mutagen 5-bromo 2-deoxyuridine and 500 ul T4 phage and put that in the shaker to determine if the flask will clear in about three hours.
	===7/12/13===		===7/12/13===

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Revision as of 21:29, 10 July 2013

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• iGEM 2013

Contents 1 July 1.1 7/1/13 1.2 7/3/13 1.3 7/5/13 1.4 7/8/13 1.5 7/10/13 1.6 7/12/13 1.7 7/15/13 2 August 2.1 8/2/13 2.2 8/5/13 2.3 8/7/13 2.4 8/9/13 2.5 8/12/13 2.6 8/14/13 2.7 8/16/13 2.8 8/19/13 2.9 8/21/13 2.10 8/23/13 2.11 8/26/13 2.12 8/28/13 2.13 8/30/13 3 September 3.1 9/2/13 3.2 9/4/13 3.3 9/6/13 3.4 9/9/13 3.5 9/11/13 3.6 9/13/13 3.7 9/16/13 3.8 9/18/13 3.9 9/20/13 3.10 9/23/13 3.11 9/25/13 3.12 9/27/13 3.13 9/30/13

July

7/1/13

7/3/13

7/5/13

7/8/13

Today we need to address some problems we ran into and make some plans for the future.

1. Plaque sizes aren't consistent, even though we are using the same agarose each time.
2. We haven't been able to observe plaque phenotypes staying the same through multiple generations
3. We have tried multiple liquid cultures and rounds of mutagenesis but we haven't been able to observe clearage in a flask yet.

Good things to model--
- Change in plaque size vs. agarose concentration
- Mutant vs wild type phage under UV - graph survival rates

7/10/13

Today we are trying out a different but similar protocol. We are hoping to see clearage in our overnight cultures which we haven't seen so far. This new protocol describes that the phage will super infect the bacteria this would explain why our phage wasn't clearing the flask. The paper states,

"After one round of infection and lysis,all remaining cells become infected. Some of these

lyse on schedule, liberating phage which superinfect the majority of cells which are still unlysed. This

superinfection causes inhibition of lysis (9). Starting at 150 min, samples (5 ml) are removed periodically

and shaken with chloroform to test for the degree of lysis which can be induced. When that test shows that

most of the cells can be induced to lyse so that clearing is nearly complete (usually at about 180 min

after initial infection),the culture is chilled in an ice bath and the pregnant bacteria are pelleted by

centrifugation at about 5,000 rpm for 10 min in a cold centrifuge."

So today we added 100ul of adenine and 200ul uricil to 25ml flasks of e.coli B in LB and in M9+ minimal media along with 500 ul mutagen 5-bromo 2-deoxyuridine and 500 ul T4 phage and put that in the shaker to determine if the flask will clear in about three hours.

7/12/13

7/15/13

August

8/2/13

8/5/13

8/7/13

8/9/13

8/12/13

8/14/13

8/16/13

8/19/13

8/21/13

8/23/13

8/26/13

8/28/13

8/30/13

September

9/2/13

9/4/13

9/6/13

9/9/13

9/11/13

9/13/13

9/16/13

9/18/13

9/20/13

9/23/13

9/25/13

9/27/13

9/30/13