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 Tuesday 1st July 1.1 Lab Work 1.1.1 Biobrick characterization 1.1.1.1 Buffer preparations 1.1.2 Visualization 1.1.2.1 Migration of plasmids containing gBlocks digestion Tuesday 1st July Lab Work Biobrick characterization Buffer preparations By Charlene and Caroline Buffer Z (250mL) Buffer Luc (250mL) Buffer STOP (15mL) Na2HPO4 – 3,125g NaH2PO4.H2O – 1,38g KCl – 0,185g MgSO4 – 0,04g β-mercaptoethanol – 0,893mL Trisphosphate pH 7,8 1M – 6,25mL MgCl2 2M – 1mL DTT 1M -0,25mL EDTA 0,5M – 0,5mL BSA – 0,25g Glycerol 60% - 62,5mL Triton 10% (100x) – 25mL Na2CO3 1M – 0,78g Visualization Migration of plasmids containing gBlocks digestion By Léa and Alice Products of the digestion made the 30/06/16 were migrated on a gel (0.8% agarose) during 30 min, 100 V. Component Volume (µL) Digested DNA 10 Loading buffer 2 For strains containing plasmids with gBlocks 4.1, according to the digestion products, 2 clones had the plasmid with the good insert. Indeed we could observe on the gel, the two fragments expected (2.7 and 0.7 Kb). All the same for strains containing gBlocks 4.2 and GFP 1-9, 3 and 2 clones respectively were good. However, DNA concentration is too low. For others strains, migration products did not get consistent results, probably due to errors during the extraction.
By Charlene and Caroline
Na2HPO4 – 3,125g
NaH2PO4.H2O – 1,38g
KCl – 0,185g
MgSO4 – 0,04g
β-mercaptoethanol – 0,893mL
Trisphosphate pH 7,8 1M – 6,25mL
MgCl2 2M – 1mL
DTT 1M -0,25mL
EDTA 0,5M – 0,5mL
BSA – 0,25g
Glycerol 60% - 62,5mL
Triton 10% (100x) – 25mL
Na2CO3 1M – 0,78g
By Léa and Alice
Products of the digestion made the 30/06/16 were migrated on a gel (0.8% agarose) during 30 min, 100 V.
For strains containing plasmids with gBlocks 4.1, according to the digestion products, 2 clones had the plasmid with the good insert. Indeed we could observe on the gel, the two fragments expected (2.7 and 0.7 Kb). All the same for strains containing gBlocks 4.2 and GFP 1-9, 3 and 2 clones respectively were good. However, DNA concentration is too low. For others strains, migration products did not get consistent results, probably due to errors during the extraction.
