Difference between revisions of "Team:Paris Saclay/Notebook/July/4"

(Digestion of plasmids)
(Digestion of plasmids)
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''By Mathilde and Alice''
 
''By Mathilde and Alice''
  
The following plasmids were digested again with EcoR1 and HindIII using [[Experiments#PlasmidDigestion|this protocol]]:
+
The following plasmids were digested again with EcoR1 and HindIII using [[Paris Saclay/Experiments#PlasmidDigestion|this protocol]]:
 
*pPS16_001 (from the 6 clones that were selected on 29/06/16)
 
*pPS16_001 (from the 6 clones that were selected on 29/06/16)
 
*pPS16_003 (from the 6 clones that were selected on 29/06/16)
 
*pPS16_003 (from the 6 clones that were selected on 29/06/16)

Revision as of 14:46, 7 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 using this protocol:

  • pPS16_001 (from the 6 clones that were selected on 29/06/16)
  • pPS16_003 (from the 6 clones that were selected on 29/06/16)
  • pPS16_005 (from the 6 clones that were selected on 29/06/16)
  • pPS16_006 (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

Plasmids extraction

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