====Transformation of DH5a cells with pPS16_020====
====Transformation of DH5a cells with pPS16_020====
Line 108:
Line 108:
''By Laetitia''
''By Laetitia''
−
Phusion PCR was performed on ligation product 2 (gblock 2.1 ligated with gblock 2.2) and ligation product 3 (gblock 3.1 ligated with gblock 3.2) made on 09/08/16.
+
Phusion PCR was performed on the ligation product 2 (gBlock 2.1 ligated with gBblock 2.2) and the ligation product 3 (gBlock 3.1 ligated with gBlock 3.2) made on 09/08/16.
It was done following [[Team:Paris_Saclay/Experiments#phusionPCR|this protocol]].
It was done following [[Team:Paris_Saclay/Experiments#phusionPCR|this protocol]].
For the PCR on ligation 2 the primers used were IPS 123 and IPS 84.
For the PCR on ligation 2 the primers used were IPS 123 and IPS 84.
Line 115:
Line 115:
Gel1
Gel1
−
====DreamTaq PCR of transformed cell with pPS16_002,008,010, 011, 012, 013, 014 and puC 19====
+
====DreamTaq PCR of transformed cell with pPS16_002,008,010, 011, 012, 013, 014 and puC19====
''By Léa and Laetitia''
''By Léa and Laetitia''
PCR DreamTaq was performed on 6 clones of each transformed cells from 10/08/16.
PCR DreamTaq was performed on 6 clones of each transformed cells from 10/08/16.
−
Each clone was plated and put in liquid culture with LB and Ampicillin.
+
Each clone was spread and put in liquid culture with LB and Ampicillin.
No PCR products were observed on the gel.
No PCR products were observed on the gel.
Line 125:
Line 125:
''By Alice''
''By Alice''
−
PSB1C3 was amplified with Phusion DNA polymerase following [[Team:Paris_Saclay/Experiments#phusionPCR|this protocol]]. Two primers ([[Team:Paris_Saclay/Experiments#primers|iPS41 and iPS42]]) were chosen. Annealing temperature was 70.9°C. Elongation step lasted 1min. After amplification, 5 µL of each PCR products with loading dye and 10 µL of DNA ladder were placed in wells and migrated at 100v during 30 min.
+
pSB1C3 was amplified with Phusion polymerase following [[Team:Paris_Saclay/Experiments#phusionPCR|this protocol]]. Two primers ([[Team:Paris_Saclay/Experiments#primers|iPS41 and iPS42]]) were used. Annealing temperature was 70.9°C. After amplification, 5 µL of each PCR products with loading dye and 10 µL of DNA ladder were placed on gel and migrated at 100v during 30 min.
PCR products expected:
PCR products expected:
Line 143:
Line 143:
''By Caroline''
''By Caroline''
−
We made the PCR again as the previous result wasn't satisfying.
+
We made the PCR again as the previous result was not satisfying.
Ligation product were cultivate overnight.
Ligation product were cultivate overnight.
[[File:T--Paris_Saclay--extraction_20160811_pcr_ligation2-3.jpg|500px|thumb|right|Result of the ligation 2-3 PCR]]
[[File:T--Paris_Saclay--extraction_20160811_pcr_ligation2-3.jpg|500px|thumb|right|Result of the ligation 2-3 PCR]]
