Line 60:
Line 60: *DS-ST1
*DS-ST1
*DS-TDCasN
*DS-TDCasN
− Then plasmids were digested with AvrII increasing the DNA quantity (10µL DNA, 1µL AvrII, 2µL Tango buffer, 7µL water). Digestion products were migrated (5µL digestion, 5µL water, 2µL {{Font color||Yellow|bleu de charge}}
+ Then plasmids were digested with AvrII
+
+ {| class="wikitable"
+ |-
+ ! Component
+ ! Volume (µL)
+ |-
+ | Plasmid
+ | 10
+ |-
+ | Tango buffer 10x
+ | 2
+ |-
+ | Water
+ | 7
+ |-
+ | AvrII enzyme
+ | 1
+ |}
+
+ The mix was incubated at 37°C for 1 hour .
+ An electrophoresis was done (0.8% of agarose).
+
+ {| class="wikitable"
+ |-
+ ! Component
+ ! Volume (µL)
+ |-
+ | Digested DNA
+ | 5
+ |-
+ | Water
+ |5
+ |
+ | Loading buffer
+ | 2
+ |}
+
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{{Team:Paris_Saclay/notebook_footer}}
Revision as of 14:52, 6 July 2016
Monday 4th July Lab work Visualization Digestion of plasmids By Mathilde and Alice
The following plasmids were digested again with EcoR1 and HindIII:
1.1 (from the 6 clones that were selected on 29/06/16)
2.1 (from the 6 clones that were selected on 29/06/16)
3.1 (from the 6 clones that were selected on 29/06/16)
3.2 (from the 6 clones that were selected on 29/06/16)
Component
Volume (µL)
Plasmid
10
Red buffer 10x
2
Water
7
EcoRI enzyme
0.5
HindIII enzyme
0.5
The mix was incubated at 37°C for 1 hour.
An electrophoresis was done (0.8% of agarose).
Component
Volume (µL)
Digested DNA
20
Loading buffer
3.3
The DNA concentration was still insufficient meaning the extraction step was not efficient.
Bringing DNA closer By Naiane and Laetitia
Plasmids from the following strains were extracted with the kit "Charge Switch-Pro Plasmid Miniprep":
DS-NMCas
DS-SPCasN
DS-ST1
DS-TDCasN Then plasmids were digested with AvrII
Component
Volume (µL)
Plasmid
10
Tango buffer 10x
2
Water
7
AvrII enzyme
1
The mix was incubated at 37°C for 1 hour.
An electrophoresis was done (0.8% of agarose).
Component
Volume (µL)
Digested DNA
5
Water
5
Loading buffer
2
