INSA-Lyon/1 October 2010
From 2010.igem.org
October 1st
Blue cells have grown, petri plate stored at room temperature.
PCR PSB1 program on plasmid extraction from 2010/09/23, with new dNTP Roche received on 2010/09/29. Protocol is the same as one from 2010/09/23. Vial 1 corresponded to the negative control with 10µl sterile water ; vial 2 corresponded to the plasmid Amp ; vial 3 corresponded to the plasmid Tet and vial 4 corresponded to the plasmid Cm.
Gel verification (0,7%): OK for the sizes of the plasmids Tet et Cm! Control negative is also OK. And PCR on plasmid Amp doesn’t produce a strip of good size (as the last time).
Clean-Up on PCR product: vial 3 and 4, and also ompR234 2010/09/20 two PCR products (kit with mini-column MN). 4 elutions are finally stored at -20°C.
DNA extraction and digestion with E of the ten clones of 1F/24A/14K. Agarose gel electrophoris : the band are not clear. no conclusion possible.
DNA extraction and digestion with E of I0500, 4O, 10L, 8C and Curli. Agarose gel electrophoris : Nothing for 8C, curli and I0500, we don't know why. 4O and 10L OK.
New DNA extraction of 4O, L2/1N, 14K, phasin, 24A, 10L and I0500. Purification on gel for 4O, L2/1N and 10L. 24A doesnt present a clear band and 14K is not visible enough. Nothing fro I0500 and phasin.
Ligation overnight at 12°C of 4O/10L and L2/1N and 18A.
liquid culture of curli, 1F/24A and 6 new clones of 1F/24A/14K
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