Toggle navigation Home Team Team Media Collaborations Sponsors Acknowledgements Project Background Design CRISPR/Cas9 Strategy Experiments Notebook Results Perspective Interlab Study Parts Parts Basic Parts Composite Parts Human Pratices Overview Societal Issues of CRISPR/Cas9 Responsible Research and Innovation GMO regulation Integrated Practices Engagement Model Attributions Safety Contents 1 Wednesday 17th August 1.1 Lab work 1.1.1 Visualization 1.1.1.1 Extraction of pPS16003, pPS16004 and GFP1-9_PSB1C3 (clones 2 and 3) 1.1.1.2 Purification of gBlocks 2, 3 and 4 1.1.1.3 Cloning with pJET the gBlock 1.2, FRB, FKBP and SgNM 1.1.1.4 Culture of BL21 Wednesday 17th August Lab work Visualization Extraction of pPS16003, pPS16004 and GFP1-9_PSB1C3 (clones 2 and 3) By Charlène Clones 2 and 3 of GFP1-9_PSB1C3, and pPS16003, pPS16004 were extracted with the Plasmid MiniPrep kit. The extracts were put to migrated on a 0.8%agarose gel with BET. Purification of gBlocks 2, 3 and 4 By Terrence The purification was carried out following the usual protocol. Cloning with pJET the gBlock 1.2, FRB, FKBP and SgNM By Léa and Naiane The cloning was carried out using a new protocol which uses pJET as cloning vector. A heat shock transformation was made on the cloning samples using the following protocol Culture of BL21 By Charlène 3 clones from each transformation condition were grown in 500µL of LB, streptomycin (50µg/mL) and chloramphenicol (30µg/mL). Then each solution was split into 3x150µL and 350µL of mix of LB + streptomycin (50µg/mL) + chloramphenicol (30µg/mL) were added to each culture. Salicylate (main solution at 10mM) was added to obtain the following concentrations : Plasmid(s) K1372001 K1372001 + pcl_TAA K1372001 + pcl_TAG K1372001 + pcl_Tq Clone 1 2 3 1 2 3 1 2 3 1 2 3 1 2 3 1 2 3 1 2 3 1 2 3 1 2 3 1 2 3 1 2 3 1 2 3 Salicylate concentration 0 30µM 1mM 0 30µM 1mM 0 30µM 1mM 0 30µM 1mM
By Charlène
Clones 2 and 3 of GFP1-9_PSB1C3, and pPS16003, pPS16004 were extracted with the Plasmid MiniPrep kit.
The extracts were put to migrated on a 0.8%agarose gel with BET.
By Terrence
The purification was carried out following the usual protocol.
By Léa and Naiane
The cloning was carried out using a new protocol which uses pJET as cloning vector.
A heat shock transformation was made on the cloning samples using the following protocol
3 clones from each transformation condition were grown in 500µL of LB, streptomycin (50µg/mL) and chloramphenicol (30µg/mL). Then each solution was split into 3x150µL and 350µL of mix of LB + streptomycin (50µg/mL) + chloramphenicol (30µg/mL) were added to each culture. Salicylate (main solution at 10mM) was added to obtain the following concentrations :
