Difference between revisions of "Haynes Lab:Notebook/Characterizing AHL quorum sensing homologs/2013/10/21"

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(mm/dd/yyyy)
(mm/dd/yyyy)
Line 7: Line 7:
 
<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->
 
<!-- ##### DO NOT edit above this line unless you know what you are doing. ##### -->
 
==mm/dd/yyyy==
 
==mm/dd/yyyy==
Retried
+
 
 +
Retry ligation from 10/19/13 '''again''' with gel extracted J06702
 +
<br>
 +
Backbone: J06702 E/X: 44 ng/ul<br>
 +
:Cut, gel purified
 +
Insert: Gblck pLac-RBS E/S: 2.6ng/ul
 +
:Cut, column purified<br>
 +
''same calculations from before, still only actually tried 3:1 and bb ctrl''<br>
 +
{| {{table}}
 +
|-
 +
| ||1:1||2:1||3:1||Bb ctrl
 +
|-
 +
|insert||0.5||1.0||1.5||0
 +
|-
 +
|vector||1.13||1.13||1.13||1.13
 +
|-
 +
|2x Roche ligation buffer||5||5||5||5
 +
|-
 +
|T4 ligase||1||1||1||1
 +
|-
 +
|H20||2.37||1.87||1.37||2.87
 +
|-
 +
|}
 +
<br>
 +
ligations ran for 3 hours at RT
 +
<br>
 +
Incubate with 50ul DH5αT for 45 min on ice
 +
<br>
 +
heat shock for 30s at 42°C
 +
<br>
 +
ice for 2min
 +
<br>
 +
spread
 +
 
 +
<!-- ##### DO NOT edit below this line unless you know what you are doing. ##### -->
 +
|}
 +
 
 +
__NOTOC__
  
  

Revision as of 17:18, 21 October 2013

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mm/dd/yyyy

Retry ligation from 10/19/13 again with gel extracted J06702
Backbone: J06702 E/X: 44 ng/ul

Cut, gel purified

Insert: Gblck pLac-RBS E/S: 2.6ng/ul

Cut, column purified

same calculations from before, still only actually tried 3:1 and bb ctrl

1:1 2:1 3:1 Bb ctrl
insert 0.5 1.0 1.5 0
vector 1.13 1.13 1.13 1.13
2x Roche ligation buffer 5 5 5 5
T4 ligase 1 1 1 1
H20 2.37 1.87 1.37 2.87


ligations ran for 3 hours at RT
Incubate with 50ul DH5αT for 45 min on ice
heat shock for 30s at 42°C
ice for 2min
spread



|}