Team:UPO-Sevilla/Notebook/07 19


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July, 19th

Production Team

Paola Gallardo. An electrophoresis was run with the products of the PCR reaction. We observed the results and came to the conclusion that the only samples that could be considered positives were two of gltD**, two of fecI-fecR*, one of fecI-fecR*-PfecA, and one of fecR*-PfecA. We were not sure if the other samples were correct, so more test had to be done. We set up inocula and spread plates of the positive samples, the first one to be used the next day and the second one to be conserved.

David Caballero. Digestion and ligation of the parts which were not confirmed in colonies PCR reactions, and that we had purified before: fecI, PfecA and fecI-fecR*. We used chloramphenicol (Cm) resistant plasmids cut previously by EcoRI and PstI (for fecI y PfecPG), and by EcoRI and SpeI (for fecI-fecR*), due to the presence of PstI target in fecI-fecR* part. We performed a PCR reaction to obtain fecA* and gltB, the biggest parts we synthesized by PCR. It was necessary to increase the elongation time to get them (from 4 to 6 minutes).

Assembly Team

Colony PCR of UPO16+3. After that, we ran an electrophoresis to see the results and there were two colonies which probably had the right plasmid. And again we digest, ligate and transformate UPO 1+19 and UPO2+13. We transformed bacteria and spread in plates. They have been growing over night.

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