IGEM:University of Illinois Urbana Champaign/2009/Notebook/Bioware 2010 RNA Decoder/2010/07/22
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July 22, 2010
Today we ran 2 digestions: one ligation for the decoder and another that is a part of coliroid. The digestions were:
The digestions were then incubated at 37 C for 15 minutes, and transfered to 80 C for 20 minutes after that. After the incubations I began ligations of the digestions.
The ligations were mixed together by flicking the tubes they were in.
The 2 ligations were incubated in room temperature (25 C) for 10 minutes and then incubated in a 80 C bath for 20 minutes.
Transformation: I did the transformations by electroporation of NEB E.coli cells that were competent. I used 50 uL of NEB cells and 3 uL of ligation mixture. I did two different methods: in one method I mixed the ligation solution in the electro-cuvette, and the other I mixed them in a PCR tube then put them into the electro-cuvette. After electroporation, I added 1 mL LB broth (SOC Outgrowth Medium) and incubated them in the 37 C room for 1 hour. Then I took 200 uL of the broth and plated onto CAP plates. So overall I made 4 plates: 2 with ligation A, and 2 with ligation B. I put the electro-cuvettes in the 4 C room.