IGEM:UNAM-Genomics Mexico/2009/Notebook/H2 Pae/2011/06/22

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Doing Competent cells

  • We centrifugate the cells which we left yesterday on CaCl and we eliminated the supernatant.
  • We added glycerol, and we divided all the cells into 20 ependorf tubes, containing 50 micro litters each.
  • With this the competent cells of E. Coli BL21_DE3 were ready.

Transformation

With the cells previously created I did the transformation of the plasmid pBBRMCS-5, which was isolated by Pato from an E. coli DH5-alfa. The transformation was done accordanly this protocol, but with just 1 tube of 50 microliters and no control, this because Miguel told me to do it in that way.

At nigth I came to see if colonies had grown, and there were around 25 colonies so Pato incubated 4 colonies in 5 ml of liquid medium(Gm) each of them, in order to isolate pBBRMCS-5 tomorrow.