Team:UNICAMP-Brazil/Notebooks/October 12

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ColiGuard

finO-biofusion - transformation results

  • Only two colonies grew in yesterday's plates.
  • Although we had experienced a low efficiency in biofusion usage, we won't continue finO's biobrick construction.

Marcelo


finOP - Final Conclusion

  • After several unsuccessful attempts on constructing finOP's biobricks, using different strategies with different backbones, our team decided to leave aside those biobricks.
  • Finalizing these parts will demand a lot of time, and we intend to already send our assembled biobricks this Thursday.
  • It's quite possible to fully assemble them, but we haven't enough time right now. We got a lot of work to do related on characterizing our assembled biobricks and updating this wiki you are currently reading on.
  • Thus, we will now focus on other kind of work. We are done with finOP inhibition system.

Marcelo


YeastGuard

New strategy: pGEM

  • The Lysozyme+biofusion2 plate grew! We screened the plate by colony PCR and found two expected fragments (nº 2 and 9). We did miniprep of these colonies.

Lisozyme-biofusion-PCR.jpg

  • We did the colony PCR to find positive colonies for the other 2 final biobricks: pJEN1 and pDLD. We found 8 positive colonies of pDLD+biofusion2 =) and 3 positive ones of pJEN1+biofusion2.

PDLD-positive.jpg
PJENOK.jpg

  • We transformed Adh1+lysozyme+Biofusion in competent E. coliand plated in LB+Amp media.

  • We did miniprep of 3 colonies of JENorf-pGEM, digested it with EcoRI and SpeI in order to release the part with the EcoRI site and to connect it in biofusion1. We found no expected fragments (1873bp). We had to decided to give up on it, because it is still on pGEM and there are two more clonnings to reach the biobrick format. We are going to focus our energy on the most promising biobricks right now.

OrfGEM+pGEM.jpg

pADH1+YFP

  • Today we made the miniprep and the digestion of the plasmids containing our new briobrick (pADH1+YFP). The enzymes used were XbaI and PstI.

  • Then, we performed the purification of the bands corresponding to the size of this new biobrick pADH1+YFP. So, it is going to be sent to IGEM organization. =)

Biobrick enviado.JPG


Wesley and Sula

YEP

  • Today we did the miniprep (Protocol 2) and we digested YEP358 – β-galactosidase plasmid with XbaI/SpeI.



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