Berk2010-Conor - Revision history

Conor McClune: /* Transposase Assembly */

Conor McClune — Sun, 17 Oct 2010 07:19:42 GMT

Transposase Assembly

←Older revision		Revision as of 07:19, 17 October 2010
Line 8:		Line 8:
	===Transposase Assembly===		===Transposase Assembly===
	====Colony PCR analysis====		====Colony PCR analysis====
		+	Colony PCR failed: no bands
		+
	*171+174 (A-D):		*171+174 (A-D):
	**lanes 1-4		**lanes 1-4

Conor McClune: /* To Do: */

Conor McClune — Sun, 17 Oct 2010 02:47:52 GMT

To Do:

←Older revision		Revision as of 02:47, 17 October 2010
Line 5:		Line 5:
	==To Do:==		==To Do:==

-	~~Things to try as payload~~ (~~all from Josh~~):	+	==[[User:Conor McClune\|Conor McClune]] 22:47, 16 October 2010 (EDT)==
-	*~~pcDNA3~~ - ~~kan resistance~~ (~~which should protect against G418~~) ~~under eukaryotic promoter~~ (~~sv40 promoter~~)	+	===Transposase Assembly===
-	*~~pcmv promoter with RFP and GFP, on two different origins of replication~~ (~~3 are amp, 2 kan, all ColE1 origin~~)	+	====Colony PCR analysis====
		+	*171+174 (A-D):
		+	**lanes 1-4
		+	**expected size: 4615
		+	**good clones:
		+	*171+175 (A-B):
		+	**lanes 5-6
		+	**expected size: 4416
		+	**good clones:
		+	*170+175 (A-C):
		+	**lanes 7-9
		+	**expected size: 5265
		+	**good clones:
		+	*170+174 (A-B):
		+	**lanes 10-11
		+	**expected size: 5464
		+	**good clones:
		+	*170+173 (A-D):
		+	**lanes 12-15
		+	**expected size: 5471
		+	**good clones:

	==[[User:Conor McClune\|Conor McClune]] 18:11, 13 October 2010 (EDT)==		==[[User:Conor McClune\|Conor McClune]] 18:11, 13 October 2010 (EDT)==

Conor McClune: /* Transposase assembly */

Conor McClune — Thu, 14 Oct 2010 01:08:47 GMT

Transposase assembly

←Older revision		Revision as of 01:08, 14 October 2010
Line 38:		Line 38:
	\| 8\|\|163-17b\|\|Xho/Bgl\|\|5400\|\|1475, 3921\|\|yes\|\|		\| 8\|\|163-17b\|\|Xho/Bgl\|\|5400\|\|1475, 3921\|\|yes\|\|
	\|-		\|-
-	\| 9\|\|2142-164a-2\|\|Xho/Bgl\|\|5090\|\|3408, 1680\|\|yes\|\|yes	+	\| 9\|\|2142-164a\|\|Xho/Bgl\|\|5090\|\|3408, 1680\|\|yes\|\|yes
	\|-		\|-
	\| 10\|\|2142-164b\|\|Xho/Bgl\|\|5090\|\|3408, 1680\|\|yes\|\|		\| 10\|\|2142-164b\|\|Xho/Bgl\|\|5090\|\|3408, 1680\|\|yes\|\|

Conor McClune: /* Transposase assembly */

Conor McClune — Thu, 14 Oct 2010 01:07:03 GMT

Transposase assembly

←Older revision		Revision as of 01:07, 14 October 2010
Line 32:		Line 32:
	\| 5\|\|163-17b\|\|Xho/Bam\|\|5400\|\|4100, 1270\|\|yes\|\|yes		\| 5\|\|163-17b\|\|Xho/Bam\|\|5400\|\|4100, 1270\|\|yes\|\|yes
	\|-		\|-
-	\| 6\|\|163-~~17b~~\|\|Xho/Bam\|\|5400\|\|4100, 1270\|\|yes\|\|	+	\| 6\|\|163-17a\|\|Xho/Bam\|\|5400\|\|4100, 1270\|\|yes\|\|
	\|-		\|-
	\| 7\|\|ladder\|\|\|\|\|\|\|\|\|\|		\| 7\|\|ladder\|\|\|\|\|\|\|\|\|\|

Conor McClune: /* Transposase assembly */

Conor McClune — Thu, 14 Oct 2010 00:39:25 GMT

Transposase assembly

←Older revision		Revision as of 00:39, 14 October 2010
Line 50:		Line 50:
	\| 14\|\|2142-166b\|\|Xho/Bgl\|\|4880\|\|3200, 1680\|\|yes\|\|yes		\| 14\|\|2142-166b\|\|Xho/Bgl\|\|4880\|\|3200, 1680\|\|yes\|\|yes
	\|}		\|}
-	Gel image:	+	Gel image:<br>
-	[[image:~~ConorGel104~~.jpg\|400px]]	+	[[image:ConorGel105.jpg\|400px]]
		+
	===preparation for plasmid payload cytometry===		===preparation for plasmid payload cytometry===
	====Self lysis test====		====Self lysis test====

Conor McClune: /* preparation for plasmid payload cytometry */

Conor McClune — Thu, 14 Oct 2010 00:33:41 GMT

preparation for plasmid payload cytometry

←Older revision		Revision as of 00:33, 14 October 2010
Line 11:		Line 11:
	==[[User:Conor McClune\|Conor McClune]] 18:11, 13 October 2010 (EDT)==		==[[User:Conor McClune\|Conor McClune]] 18:11, 13 October 2010 (EDT)==

		+	===Transposase assembly===
		+	Conducted the following digestions:
		+	{\| {{table}}
		+	\| align="center" style="background:#f0f0f0;"\|'''Lane'''
		+	\| align="center" style="background:#f0f0f0;"\|'''Part'''
		+	\| align="center" style="background:#f0f0f0;"\|'''Restr. Enzymes'''
		+	\| align="center" style="background:#f0f0f0;"\|'''Parent Size'''
		+	\| align="center" style="background:#f0f0f0;"\|'''Expected Fragments'''
		+	\| align="center" style="background:#f0f0f0;"\|'''Acceptable?'''
		+	\| align="center" style="background:#f0f0f0;"\|'''Gel Purified'''
		+	\|-
		+	\| 1\|\|163-5a\|\|Xho/Bam\|\|6004\|\|4730, 1270\|\|yes\|\|
		+	\|-
		+	\| 2\|\|163-5b\|\|Xho/Bam\|\|6004\|\|4730, 1270\|\|yes\|\|yes
		+	\|-
		+	\| 3\|\|163-10a\|\|Xho/Bam\|\|5150\|\|3880, 1270\|\|no\|\|
		+	\|-
		+	\| 4\|\|163-10b\|\|Xho/Bam\|\|5150\|\|3880, 1270\|\|no\|\|
		+	\|-
		+	\| 5\|\|163-17b\|\|Xho/Bam\|\|5400\|\|4100, 1270\|\|yes\|\|yes
		+	\|-
		+	\| 6\|\|163-17b\|\|Xho/Bam\|\|5400\|\|4100, 1270\|\|yes\|\|
		+	\|-
		+	\| 7\|\|ladder\|\|\|\|\|\|\|\|\|\|
		+	\|-
		+	\| 8\|\|163-17b\|\|Xho/Bgl\|\|5400\|\|1475, 3921\|\|yes\|\|
		+	\|-
		+	\| 9\|\|2142-164a-2\|\|Xho/Bgl\|\|5090\|\|3408, 1680\|\|yes\|\|yes
		+	\|-
		+	\| 10\|\|2142-164b\|\|Xho/Bgl\|\|5090\|\|3408, 1680\|\|yes\|\|
		+	\|-
		+	\| 11\|\|2142-165a\|\|Xho/Bgl\|\|5080\|\|3401, 1680\|\|yes\|\|yes
		+	\|-
		+	\| 12\|\|2142-165b\|\|Xho/Bgl\|\|5080\|\|3401, 1680\|\|yes\|\|
		+	\|-
		+	\| 13\|\|2142-166a\|\|Xho/Bgl\|\|4880\|\|3200, 1680\|\|yes\|\|
		+	\|-
		+	\| 14\|\|2142-166b\|\|Xho/Bgl\|\|4880\|\|3200, 1680\|\|yes\|\|yes
		+	\|}
		+	Gel image:
		+	[[image:ConorGel104.jpg\|400px]]
	===preparation for plasmid payload cytometry===		===preparation for plasmid payload cytometry===
	====Self lysis test====		====Self lysis test====

Conor McClune: /* Conor McClune 18:11, 13 October 2010 (EDT) */

Conor McClune — Wed, 13 Oct 2010 23:43:51 GMT

Conor McClune 18:11, 13 October 2010 (EDT)

←Older revision		Revision as of 23:43, 13 October 2010
Line 28:		Line 28:
	*added 40 uL bacteria to each test tube at 2:30		*added 40 uL bacteria to each test tube at 2:30
	*induced with arabinose fifteen minutes later (at 2:45)		*induced with arabinose fifteen minutes later (at 2:45)
		+
	==[[User:Conor McClune\|Conor McClune]] 18:31, 12 October 2010 (EDT)==		==[[User:Conor McClune\|Conor McClune]] 18:31, 12 October 2010 (EDT)==
	*flow cytometry got no hits, because the choanos were in ASW, so the bacteria did not lyse		*flow cytometry got no hits, because the choanos were in ASW, so the bacteria did not lyse

Conor McClune at 23:33, 13 October 2010

Conor McClune — Wed, 13 Oct 2010 23:33:40 GMT

←Older revision		Revision as of 23:33, 13 October 2010
Line 14:		Line 14:
	====Self lysis test====		====Self lysis test====
	*started 2:00		*started 2:00
-	*checked at ~~____~~:	+	*checked at 4:30:
	{\| {{table}}		{\| {{table}}
	\| \|\|1\|\|2\|\|3\|\|6\|\|7		\| \|\|1\|\|2\|\|3\|\|6\|\|7
	\|-		\|-
-	\| 51\|\|\|\|\|\|\|\|\|\|	+	\| 51\|\|lysed\|\|lysed\|\|lysed\|\|barely lysed\|\|lysed
	\|-		\|-
-	\| 51\|\|\|\|\|\|\|\|\|\|	+	\| 51\|\|lysed\|\|lysed\|\|lysed\|\|lysed\|\|lysed
	\|}		\|}

Conor McClune: /* Conor McClune 18:31, 11 October 2010 (EDT) */

Conor McClune — Wed, 13 Oct 2010 22:11:44 GMT

Conor McClune 18:31, 11 October 2010 (EDT)

←Older revision		Revision as of 22:11, 13 October 2010
Line 8:		Line 8:
	*pcDNA3 - kan resistance (which should protect against G418) under eukaryotic promoter (sv40 promoter)		*pcDNA3 - kan resistance (which should protect against G418) under eukaryotic promoter (sv40 promoter)
	*pcmv promoter with RFP and GFP, on two different origins of replication (3 are amp, 2 kan, all ColE1 origin)		*pcmv promoter with RFP and GFP, on two different origins of replication (3 are amp, 2 kan, all ColE1 origin)
		+
		+	==[[User:Conor McClune\|Conor McClune]] 18:11, 13 October 2010 (EDT)==
		+
		+	===preparation for plasmid payload cytometry===
		+	====Self lysis test====
		+	*started 2:00
		+	*checked at ____:
		+	{\| {{table}}
		+	\| \|\|1\|\|2\|\|3\|\|6\|\|7
		+	\|-
		+	\| 51\|\|\|\|\|\|\|\|\|\|
		+	\|-
		+	\| 51\|\|\|\|\|\|\|\|\|\|
		+	\|}
		+
		+
		+	====preparation of cells====
		+	*scraped choanos and put 1ml in each test tube
		+	*added 40 uL bacteria to each test tube at 2:30
		+	*induced with arabinose fifteen minutes later (at 2:45)
		+	==[[User:Conor McClune\|Conor McClune]] 18:31, 12 October 2010 (EDT)==
		+	*flow cytometry got no hits, because the choanos were in ASW, so the bacteria did not lyse

	==[[User:Conor McClune\|Conor McClune]] 18:31, 11 October 2010 (EDT)==		==[[User:Conor McClune\|Conor McClune]] 18:31, 11 October 2010 (EDT)==

Conor McClune: /* preparation for plasmid payload cytometry */

Conor McClune — Mon, 11 Oct 2010 23:19:07 GMT

preparation for plasmid payload cytometry

←Older revision		Revision as of 23:19, 11 October 2010
Line 13:		Line 13:
	====Self lysis test====		====Self lysis test====
	*started 2:15		*started 2:15
		+	*checked at 4:15:
		+	{\| {{table}}
		+	\| \|\|1\|\|2\|\|3\|\|6\|\|7
		+	\|-
		+	\| 51\|\|lysed\|\|lysed\|\|lysed\|\|lysed\|\|lysed
		+	\|-
		+	\| 51\|\|lysed\|\|lysed\|\|lysed\|\|lysed\|\|lysed
		+	\|}