Team:UNICAMP-Brazil/Notebooks/October 3

From 2009.igem.org

(Difference between revisions)
(Transformation of finOP and Cre-Recombinase's ligations)
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''Marcelo and Victor''
''Marcelo and Victor''
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==''' YeastGuard '''==
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====New biobricks - New strategy (pGEM)====
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* The ligation reactions didn´t work. So today we decided not to digest the fragment and the pGEM vector with ''SpeI'', but to ligate it directly, considering that we can confirm the correct insertion by PCR.
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''Taís''
{{:Team:UNICAMP-Brazil/inc_rodape}}
{{:Team:UNICAMP-Brazil/inc_rodape}}

Revision as of 05:22, 16 October 2009

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ColiGuard

Ligation of finOP and Cre-Recombinase on pGEM vector

  • In order to start our pGEM strategy on assembling biobricks, today we ligated PCR's products of finO, finP and Cre-Recombinase into pGEM Vector. There is no need for any digestion before this procedure, since pGEM has an T stick ends capable of pairing with the A stick ends left by Taq Polymerase on the PCR products.
  • We followed protocol 11, without modifications.
  • Ligation lasted 1 hour.

Marcelo and Victor

Transformation of finOP's and Cre-Recombinase's ligations

  • After performing the required ligations, we then transformed them into electrocompetent E. coli bacteria, strain DH10B, according to protocol 3.
  • We plated the trasformed cells in LB-AMP media, which already contained X-gal substrate.
  • Plates were incubated at 37ºC for an O/N period.

Marcelo and Victor

YeastGuard

New biobricks - New strategy (pGEM)

  • The ligation reactions didn´t work. So today we decided not to digest the fragment and the pGEM vector with SpeI, but to ligate it directly, considering that we can confirm the correct insertion by PCR.

Taís