Revision as of 18:53, 16 September 2008

Purification of yesterday digestion

D112 before gel excision

Digestion n°	DNA substrat	Digestion by
D112	MP106: S03879 (rbs-TetR) in pSB1A2	EcoRI & SpeI

1% agarose gel

Qiaquick Gel Extraction Kit

Sample	DNA concentration	A260/A280
D112	8 µg/mL	2,31

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 57: / Line 57: @@
 *L184 = pFliL - ECFP LVA+
 *L185 = pFliL - ECFP LVA-
+==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