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 Friday 12th August 1.1 Visualization 1.1.1 PCR of pPS16_020 1.1.2 Extraction of pUC19 1.1.3 Dreamtaq PCR on puc19, detection, FRB, FKBP, SgRNAst, SgRNAnm and gBlock 1.2 Friday 12th August Visualization PCR of pPS16_020 By Léa A Dreamtaq PCR was performed on pPS16_009 using the usual protocol. Primers: IPS83 ans IPS84 Electrophoresis of PCR products, using primers IPS 83 and IPS 84 on pPS16_016. Extraction of pUC19 By Charlène Clones 1 and 2 of pUC19 were extracted with the Plasmid MiniPrep kit. They were fast digested with EcoRI : 3µL of plasmid + 1µL of FD buffer + 0.5µL of FD EcoRI + 5.5µL of water, 15 minutes at 37°C. pUC19 clones 1 and 2 digested by EcoRI Just clone 1 is at the good size so we will have to use it and not clone 2. Dreamtaq PCR on puc19, detection, FRB, FKBP, SgRNAst, SgRNAnm and gBlock 1.2 By Naiane, Mahnaz and Terrence A Dreamtaq PCR was performed on puc19, detection, FRB, FKBP, SgRNAst, SgRNAnm and gBlock 1.2 using the usual protocol with these volumes : Components Volume 10X DreamTaq Green Buffer 2.5µL dNTP (10mM) 1µL Primers mix (10µM each) 1µL DreamTaq DNA polymerase 0.25µL Nuclease-free water up to 25µL Total volume 25µL We made 6 clones of each colonie except for Puc19 where we made 1 negative control. Result of the PCR : there is no band at the expected size for all the screened clones so the transformations of the gblock do not seem to have worked. 1.2 and FRB Detection and ST puc19, NM and FKBP FKBP
By Léa
A Dreamtaq PCR was performed on pPS16_009 using the usual protocol. Primers: IPS83 ans IPS84
By Charlène
Clones 1 and 2 of pUC19 were extracted with the Plasmid MiniPrep kit.
They were fast digested with EcoRI : 3µL of plasmid + 1µL of FD buffer + 0.5µL of FD EcoRI + 5.5µL of water, 15 minutes at 37°C.
Just clone 1 is at the good size so we will have to use it and not clone 2.
By Naiane, Mahnaz and Terrence
A Dreamtaq PCR was performed on puc19, detection, FRB, FKBP, SgRNAst, SgRNAnm and gBlock 1.2 using the usual protocol with these volumes :
We made 6 clones of each colonie except for Puc19 where we made 1 negative control.
Result of the PCR : there is no band at the expected size for all the screened clones so the transformations of the gblock do not seem to have worked.
