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• Successful assembly of parts [http://partsregistry.org/wiki/index.php?title=Part:BBa_K175023 K175023] and [http://partsregistry.org/wiki/index.php?title=Part:BBa_K175024 K175024] which correspond to Plasmid 1 and 2 of the Biosynthetic AND gate section respectively.
• Electroporation of plasmid 1 and 2 into Escherichia coli TOP10 cells.

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The assembly of the plasmid 1 ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K175023 K175023]) and 2 ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K175024 K175024]) was successful. As it can be noticed in the gel electrophoresis image, the bands are in agreement with the expected sizes (Figure E). After the last assembly step (it is worth to mention that all assemblies were carried using the biobrick assembly kit), 2µL of the resulted DNA mix was electroporated into Escherichia coli TOP10 cells. This resulted in two 1.5 mL eppendorf tubes containing cells with either plasmid 1 or plasmid 2.

250µL of cells carrying either plasmid 1 or 2 were used to inoculate one LB agar plate containing the proper antibiotic as selection marker. The plates were incubated at 37ºC overnight. One of the resulted colonies from each plate were used to inoculate one 5mL LB tube each. The tube was incubated overnight at 37ºC and 175 rpm. The resulted cultures were used to obtain miniprep plasmid DNA which was sent to sequencing. Unfortunately, the sequence showed no match with any of the planed biobricks. As the time was running and less people could be in charge of the lab work, this strategy was left behind and the efforts were directed to the lock/key library and the synthetic transcriptional cascade.

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