August, 9th

Production Team

David Caballero. We analyzed plaques spread the day before; it looked like if the bacteria could not grow. There were only three red colonies (negatives) in a plaque. Those devices had promoter and were assembled in high copy vectors. This is why we thought that the high expression of the devices could be harmful for bacteria. We analyzed the digestions in agarose gel to verify it. Al most all the digestions were well made and we only had to repeat two.

In order to avoid that fact again, we assembled devices including promoters in low copy vectors. Like in before cases, we used different vectors depending of the resistance of the origin vectors we need. It is summed up down:

• pSB4K5: (1+2) + (4)
• pSB4C5: (1+2) + (7+2)
• pSB3T5: (11) + (19) // (12) + (19)
• pSB3C5: (11+2) + (13+3) // (12+2) + (13+3) // (11+2) + (16+3) // (12+2) + (16+3) // (1+2) + (16+3)

It was made the necessary digestions and we left ligation over night at 13ºC on average.

Assembly Team

Digestion: 2+5+3 with BamI-PstI. We started the following devices (3A method): 7+2+28; 2+6+2+5+3; 2+18; 28+3; 2+4+2+6; 2+28; 18+3: digestion, ligation and transformation.