Line 59:
Line 59: Plasmids were resuspended in 100μL of Milli-Q water.
Plasmids were resuspended in 100μL of Milli-Q water.
DNA stored at -20°C.
DNA stored at -20°C.
+
+
+
+ ====pPS16_001, pPS16_002, pPS16_009, pPS16_014 DreamTaq PCR ====
+ ''By Mathilde and Laetitia''
+
+ DreamTaq PCR was carried out following the usual [[Team:Paris_Saclay/Experiments#taqPCR|protocol]].
+ Only the clone 9 for PS16_001 (1.1), the clone 11 for PS16_002 (1.2), the clone 7 for PS16_009 (GFP) and clones 7 and 11 for pPS16_014 (Nm) were right.
===Interlab Study===
===Interlab Study===
Revision as of 09:30, 22 August 2016
Contents
1 Wednesday 3rd August
1.1 Lab work
1.1.1 Visualization
1.1.1.1 Glycerol stocks 1.1.1.2 High fidelity Q5 PCR on pPS16_004, pPS16_006 and pPS16_007 1.1.1.3 Extraction of the plasmids containing gBlocks pPS16_001, pPS16_002, pPS16_003, FRB, sg-ST1, FKPB, DS-NMcasN-, spacer and detection 1.1.1.4 Extraction of the plasmids containing gBlocks 1.1.1.5 pPS16_001, pPS16_002, pPS16_009, pPS16_014 DreamTaq PCR
1.1.2 Interlab Study
Wednesday 3rd August Lab work Visualization Glycerol stocks By Caroline
The bacteria transformed with the plasmids sent the 2/08/2016 and those that would be sent the 4/08/2016 were put into glycerol. 1mL of liquid culture and 0.5mL of glycerol were put at -20°C.
High fidelity Q5 PCR on pPS16_004, pPS16_006 and pPS16_007 By Caroline
Q5 PCR was carried out following the usual protocol adapted to 50µL at a Tm of 72°C. The primers used were specific to amplify only the sequences of interest. The products were put for migration on a 0.8%agarose gel with BET. Only, PCR form pPS16_004 and pPS16_007 showed a positive result.
Extraction of the plasmids containing gBlocks pPS16_001, pPS16_002, pPS16_003, FRB, sg-ST1, FKPB, DS-NMcasN-, spacer and detection By Laetitia
The extraction was performed without kit, following the usual protocol .
It was done on:
Puc 19 containing
Clone
pPS16_001
4
pPS16_002
3
pPS16_003
2, 5
FRB
5
sg-ST1
2, 3, 5
DS-NMcasN-
3
FKBP
4
Spacer
1, 6
Detection
3, 4, 6
By Naiane
The kit "Charge Switch-Pro Plasmid Miniprep" was used to extract plasmidic DNA of the plasmid pPS16_003 containing the Gblock 2.1, plasmid pPS16_004 containing the Gblock 2.2, plasmid pPS16_006 containing the Gblock 3.2 and the plasmid pPS16_007 containing the Gblock 4.1 from 3mL of overnight culture.
Plasmids were resuspended in 100μL of Milli-Q water.
DNA stored at -20°C.
pPS16_001, pPS16_002, pPS16_009, pPS16_014 DreamTaq PCR By Mathilde and Laetitia
DreamTaq PCR was carried out following the usual protocol .
Only the clone 9 for PS16_001 (1.1), the clone 11 for PS16_002 (1.2), the clone 7 for PS16_009 (GFP) and clones 7 and 11 for pPS16_014 (Nm) were right.
Interlab Study Preculture of device one transformed DH5a By Lea
Two colonies were inoculated inside of two tubes of 3mL of liquid LB medium containing 30 µg/ml Chloramphenicol. The tubes were incubated at 37°C overnight.
