Latest revision as of 18:55, 16 September 2008

Cloning of EnvZ*

Defroze of the cells on ice
Add 5 µL of DNA in 100 µL of competent cells (ligation products or pUC19 for positive control)
30 min on ice
Heat schock 45sec at 42°C
2 min on ice
Add 900 µL of SOC
Incubate 1h at 37°C
Centrifugate 5 min at 6000 rpm
Remove 800 µL
Plate the 200 µL left on LB + amplicilline
Incubate O/N at 37°C

@@ Line 7: / Line 7: @@
 {|border="1" style="text-align: center"
 |
-|DNA concentration
+|'''DNA concentration'''
-|A260 / A2801
+|'''A260 / A2801'''
 |-
 |D159 (EnvZ*)
@@ Line 23: / Line 23: @@
 {|border="1" style="text-align: center"
 |rowspan="2"|
-|rowspan="2"|control
+|rowspan="2"|'''control'''
-|colspan="2"|insert / vector ratio
+|colspan="2"|'''insert / vector ratio'''
 |-
-|3 / 1
+|'''3 / 1'''
-|4 / 1
+|'''4 / 1'''
 |-
 |D159 (EnvZ*)
@@ Line 54: / Line 54: @@
 |3,4 µL
 |}
+*10 min at room temperature
+*followed by 7 hours at about 16°C.
+==Transformation of E. coli DH5 alpha competent cells==
+* Defroze of the cells on ice
+* Add 5 µL of DNA in 100 µL of competent cells (ligation products or pUC19 for positive control)
+* 30 min on ice
+* Heat schock 45sec at 42°C
+* 2 min on ice
+* Add 900 µL of SOC
+* Incubate 1h at 37°C
+* Centrifugate 5 min at 6000 rpm
+* Remove 800 µL
+* Plate the 200 µL left on LB + amplicilline
+* Incubate O/N at 37°C