IGEM:Virginia/2012/Notebook/Genetically engineered bacteriophage for diagnosis of whooping cough/2012/09/16

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Week of 9/16/12

Performed gel extraction of ligated backbones (both E+P and E+S); failed to show up in adequate amounts when nanodropped.
Ligated Plasmid+RBS (E+S), SynCG (X+P), and RFP (X+P) at 3:1 and 1:1 ratios of both 2X ligase buffer and promega DNA ligase and performed PCR.
Repeated ligation and performed PCR purification: P+RBS+RFP and P+RBS+SynCG demonstrate proper amounts of DNA but at low concentrations, suggesting that perhaps the DNA is not the intended amplicon.
Discovered that roughly half of the previous pregnancy tests now appear positive, brighter than evaporation lines would be. Some tests for 5 mM DTT (denaturing solution), 0.5 mM DTT, 0.05 mM DTT, and 0 mM DTT are now positive with the majority in the latter two groups.
Nanodropped the P+RBS+RFP, P+RBS+SynCG, backbone (E+S), backbone (E+P), and SynCG all (E+P): only the backbone (E+S) and SynCG all (E+P) showing proper concentrations. The latter includes the IDT plasmid backbone.
Plated the bbp9 biobricks (both 3:1 and 1:1 ratios), the hCG biobrick (in 3:1, 1:1, and 1:3 ratios), backbone, original BBa_K331022, and a DNA lacking group, producing thousands of cells on each plate (except for the last), demonstrating that many of the cells have empty vectors.